Chapter 26

BIOSAFETY

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Work Process C. Biosafety Principles and Levels

To determine which controls are required to mitigate hazards and perform work safely, supervisors and work leads must understand and apply the processes and requirements for defining work, identifying hazards, and assessing risks, as discussed in Work Process B. Controls are safeguards employed to contain biological agents or materials and therefore prevent the exposure of workers, other people, or the environment to agents that may harm them.

In biosafety, the term “containment” describes the set of controls, including safe methods, equipment, and facilities needed to protect workers and the environment from biohazardous materials or agents. Controls used for containment in laboratories are described in Biosafety in Microbiological and Biomedical Laboratories (BMBL), Section III, as the Principles of Biosafety. These containment controls are listed below and summarized in the next sections:

The Berkeley Lab Biosafety Work Authorization is used to define work, identify hazards, assess risks, and implement any of the containment controls listed above. See Work Process A and Work Process D.2 for additional information on work authorization documents.

1.    Laboratory Practices

The first and most important element of control for laboratory containment and research product protection is strict adherence to laboratory biosafety containment practices and good microbiological practice (GMP). Biosafety containment practices include standard microbiological practices and special practices specified by the CDC and NIH. GMP is based on widely accepted aseptic practices.

a.    Standard microbiological practices and special practices are administrative controls listed as BL containment criteria in BMBL and the NIH Guidelines to protect workers and the environment. (See Work Process C.4.a of this manual for additional information on BL containment categories and criteria.) These practices, along with requirements from other biosafety standards, are used for the safe performance of work documented in the Berkeley Lab Biosafety Work Authorization. Standard microbiological practices or special practices for laboratories apply to most Berkeley Lab work with biological materials. Standard practices for BL1 and BL2 laboratories address the following topics (see Appendix C and Work Process D for more information):

b.    Good microbiological practice (GMP) is also typically needed for containment and good research. GMP is based on aseptic techniques and other good microbiological practices necessary to prevent contamination of the laboratory with the agents being handled and contamination of the work with agents from the environment. See Appendix D for common GMP. Supervisors and work leads are responsible for selecting and instructing workers on the specific GMP needed to conduct the work, or additional safety practices needed for specific agents or procedures.

2.    Safety and Personal Protective Equipment

Worker exposure to infectious agents may be prevented by the use of standard and activity-specific safety and personal protective equipment (PPE) as primary barriers or controls. The need for additional activity-specific safety equipment or PPE must be determined during risk assessment, and any equipment needed for safety should be included in the Biosafety Work Authorization.

Standard safety equipment and PPE are equipment controls listed as biosafety level (BL) containment criteria in BMBL and the NIH Guidelines. They provide primary barriers that prevent worker exposure to infectious agents. See Work Process C.4.a for additional information on BL containment categories and criteria. These standard equipment requirements, along with requirements from other biosafety standards, are used and customized for the work to be conducted. They are also summarized in the Berkeley Lab Biosafety Work Authorization. Standard safety equipment and PPE are applicable to most work with biological materials at Berkeley Lab. The following types of standard equipment and PPE are further discussed in Appendix C and Work Process D:

3.    Facility Design and Construction

Facility design and equipment provide secondary barriers that protect laboratory workers, persons outside the laboratory, the public, and the environment from potentially hazardous materials or agents that may be accidentally released from the laboratory.

Picture Picture

Standard facilities provide secondary barriers.
Source: HHS CDC Office of Health and Safety.

Berkeley Lab designs, constructs, approves, and operates its buildings and laboratories in accordance with nationally recognized codes regulations, standards, acts, and manuals as detailed in the Facilities Division Construction Details and Design Guidelines and ES&H Manual’s Fire Prevention and Protection program. In addition, EHS personnel are assigned to conduct design reviews for new projects as described in the Capital Projects Procedures Manual (PUB-3193). Specifically assigned EHS personnel are responsible for reviewing facility design and installation for concerns and requirements related to biosafety. Line management must ensure that facilities are appropriate for the laboratory activities and meet the required BL for the agents or materials being handled.

In addition, Berkeley Lab designs and operates its facilities in which work with biological materials is conducted in accordance with applicable standard facilities criteria. “Standard facilities” are design features, materials, and equipment incorporated into the laboratory or facility in accordance with BL containment criteria stated in BMBL and the NIH Guidelines (see Work Process C.4, Biosafety Containment Levels and Criteria, for more information).

Standard BL1 and BL2 laboratory facility barriers are sufficient to control most work at Berkeley Lab. This is because risks related to most work are associated with direct contact with materials or agents in standard laboratories. Examples of standard facility barriers and equipment are listed below. See Appendix C for standard laboratory facilities criteria that summarize how these barriers must be employed. See Work Process D for additional information on each of these topics.

If the risk assessment indicates a risk of exposure to an infectious aerosol, then higher levels of safety equipment and PPE (primary barriers) or multiple secondary facilities barriers are necessary. Multiple secondary facilities barriers are not typically needed at Berkeley Lab.

Some standard facilities are summarized in the Biosafety Work Authorization. Any additional special facility barriers that are required should also be included in the authorization.

4.    Biosafety Containment Levels and Criteria

Berkeley Lab requires researchers who work with biological materials to implement containment controls in accordance with an established BL. BL is a standard combination of practices and techniques, safety equipment, and facilities to safely contain biohazardous materials or agents to be used in the work, as specified by BMBL or the NIH Guidelines.

Work at Berkeley Lab requires routine application of BLs developed for biological laboratories, and occasional application of BLs developed for other types of work such as large-scale recombinant operations. BLs for laboratories are presented in the next section. BLs for large-scale, plant, and animal uses are presented in Work Process C.4.b, Additional Containment Categories, of this program.

The appropriate BL must be selected once the risk assessment has been completed. The final BL determination should consider all aspects of the work, hazards, and controls. The PI or supervisor should propose the appropriate BL(s) when submitting the authorization for review. The final BL(s) are determined by the Institutional Biosafety Committee (IBC).

a.    Laboratory Containment Levels. Containment controls for laboratory biosafety are categorized into four BLs. Definitions of each laboratory BL are provided in Table 8. Work at Berkeley Lab is commonly conducted at BL1 or BL2; work at BL3 or BL4 is not currently conducted. Laboratory work at Berkeley Lab must be conducted in accordance with the standard and special work practices, safety equipment, and facility requirements noted in the laboratory BL1 and BL2 criteria listed in Appendix C.

Table 8. Laboratory Biosafety Containment Levels

Present
at Berkeley Lab

Biosafety Level (BL)

Biosafety Level Definition

Yes

1

BL1 is suitable for work involving agents of unknown or minimal potential hazard to laboratory personnel and the environment,1 or work with defined and characterized strains of viable microorganisms not known to consistently cause disease in healthy adult humans.2

2

BL2 is suitable for laboratory work involving agents of moderate potential hazard to personnel and the environment.1 Primary hazards to personnel working with these agents relate to accidental percutaneous or mucous membrane exposures, or ingestion of infectious materials.2

No

3

BL3 is applicable to facilities in which work is conducted with indigenous or exotic agents that may cause serious or potentially lethal disease as a result of exposure by the inhalation route.3

4

BL4 is required for work with dangerous and exotic agents that pose a high individual risk of life-threatening disease, aerosol transmission, or related agents having an unknown transmission risk.3

Footnotes:

  1. NIH Guidelines
  2. BMBL, fifth edition, Section III
  3. BMBL, fifth edition, Section IV

 

When developing the Biosafety Work Authorization, the appropriate laboratory BL must be selected after conducting the risk assessment. Typical BLs used for various materials and agents are listed in Work Process C.4.a.i, Laboratory Biosafety Level 1, and Work Process C.4.a.ii, Laboratory Biosafety Level 2. The final BL(s) are determined by the IBC.

                                                     i.      Laboratory Biosafety Level 1 Text Box:

BL1 laboratories are not necessarily separated from the general traffic patterns in the building. Work is typically conducted on open benchtops using standard microbiological practices. Special containment equipment or facility design is not required, but may be used as determined by a risk assessment. Laboratory personnel must have specific training in the procedures conducted in the laboratory and must be supervised by a scientist trained in microbiology or a related science.

 
Laboratory BL1 work with open benchtops and standard
microbiological practices.
Source: Berkeley Lab EHS.

 

The BL will be determined as part of the risk assessment. BL1 containment is typically required for laboratory work involving:

                                                   ii.      Laboratory Biosafety Level 2 Text Box:Text Box:

BL2 laboratories follow BL1 requirements and additional BL2 requirements such as:

Text Box:  
BL2 work inside a BSC.
Source: Berkeley Lab EHS.
BL2 work inside a BSC. Source: Berkeley Lab EHS.

The BL will be determined as part of the risk assessment. Laboratory BL2 containment is typically required for laboratory work involving:

b.    Additional Containment Categories

Additional types of containment specified in the NIH Guidelines or BMBL may also be applicable to work with biological materials at Berkeley Lab. Table 9 below lists these additional containment categories and relevant NIH Guidelines or BMBL section. When selecting a containment level for a type of work listed in Table 9, the supervisor, work lead, and IBC should carefully review this table to determine if the containment category and criteria apply to the planned Berkeley Lab work.

Laboratory- or operation-specific authorizations, biosafety manuals, or other documents may be used to document the containment requirements related to the work. If the containment categories or criteria presented in Table 9 are not applicable to the work, the laboratory BLs presented in Work Process C.4.a are applied.

                                                     i.      Text Box:Recombinant Large-Scale Containment Levels

Physical containment guidelines from Appendix K of the NIH Guidelines must be used for large-scale research or production activities involving viable organisms containing recombinant DNA molecules. “Large scale (BL–Large Scale)” is a term used in the NIH Guidelines and the Berkeley Lab biosafety policy to describe uses of and containment levels for organisms containing recombinant DNA molecules involving a quantity of culture greater than 10 liters. Note that this quantity category typically means the quantity of a material in a single batch of liquid culture; however, this batch quantity is not defined by NIH and should be used as a guideline to determine the applicability of large-scale containment criteria. Criteria for large-scale containment address the biological hazard associated with organisms containing recombinant DNA only. Large-scale containment criteria must be selected based on the findings of the risk assessment, and then documented in the Biosafety Work Authorization.

 

Large-scale processes and containment. Left: Photobioreactor process. Source: Berkeley Lab EHS.
Right: Fermentation process to produce biofuel. Source: Berkeley Lab Public Affairs.

 

                                                    ii.      Text Box:Recombinant Plant Containment Levels

Biosafety containment levels and criteria for recombinant research with plants must be selected based on the findings of the risk assessment, and then documented in the Biosafety Work Authorization. Laboratory or plant biosafety containment levels must be applied to the work as follows:

Text Box:  
Arabidopsis sp. Source: Berkeley Lab EHS.
Arabidopsis sp. Source: Berkeley Lab EHS.

Containment Category

Standard and Section

Focused Scope of Containment Criteria

Large-scale uses of organisms containing recombinant DNA molecules

NIH Guidelines, Appendix K

Physical containment guidelines for large-scale (greater than 10 liters of culture) research or production activities involving viable organisms containing recombinant DNA molecules

Recombinant DNA research involving plants

NIH Guidelines, Appendix P

Physical and biological containment conditions and practices suitable to greenhouse operations that conduct experiments involving plants, plant-associated microorganisms, and small animals (e.g., arthropods or nematodes)

Recombinant DNA research involving animals

NIH Guidelines, Appendix Q

Containment and confinement practices for research involving whole animals when the animals are of a size or have growth requirements that preclude the use of containment for laboratory animals (i.e., including but not limited to nonhuman primates, cattle, swine, sheep, goats, horses, and poultry) and:

  • The animals’ genomes have been altered by the introduction of recombinant DNA or DNA derived therefrom into the germ line (transgenic animals), or
  • Experiments involving viable recombinant DNA-modified microorganisms have been tested on whole animals.

Vertebrate animal BL criteria for indoor research facilities (e.g., vivaria)

BMBL,
Section V

Use of experimentally infected animals housed in indoor research facilities (e.g., vivaria), and the maintenance of laboratory animals that may naturally harbor zoonotic infectious agents

Arthropod containment
guidelines

BMBL, Appendix E

Risk assessment and containment for arthropods of public-health importance, including those that transmit pathogens. Arthropods that only bite, sting, or cause myiasis and infestation are not included. Myiasis is an infestation of tissue by fly larvae, or a disease resulting from such infestation.


The plant BLs listed in Appendix P of the NIH Guidelines specify physical and biological containment conditions and practices suitable for conducting greenhouse experiments involving recombinant DNA-containing plants, plant-associated microorganisms, and small animals. Acronyms for plant BLs are BL1-P through BL4-P. The following bullets further clarify terms and applicability of the plant biosafety levels:

Text Box:  
Plant containment (BL1-P) in a greenhouse facility. Source: Berkeley Lab EHS.
Plant containment (BL1-P) in a greenhouse facility. Source: Berkeley Lab EHS.

A Practical Guide to Containment developed by Virginia Polytechnic and State University is a good example of how to apply plant BLs in research with greenhouse transgenic plants and microbes.

                                                  iii.      Vertebrate Animal Containment Levels

Biosafety containment levels and criteria for the use or care of vertebrate animals must be selected or developed following the risk assessment and covered in the Biosafety Work Authorization. Laboratory or animal biosafety containment levels must be applied to the work as follows:

  

Small-animal containment at BL1 (left) and BL2 with a
BSC (right).
Source: Berkeley Lab EHS.

 

Agent summary statements for zoonotic agents in Section VIII of BMBL also recommend containment levels for laboratory use of zoonotic agents, and for handling animals infected with the agent.

                                                  iv.      Text Box:Arthropod Containment Levels

Biosafety containment levels and criteria for the use of arthropods must be selected or developed after the risk assessment. Laboratory or arthropod biosafety containment levels must be applied to the work as follows:

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