The Laboratory Biosafety Guidelines: 3rd Edition 2004

Chapter 2
Biological Safety

2.1 Risk Groups

Classification of organisms according to risk group has traditionally been used to categorize the relative hazards of infective organisms. The factors used to determine which risk group an organism falls into is based upon the particular characteristics of the organism, such as

• pathogenicity
• infectious dose
• mode of transmission
• host range
• availability of effective preventive measures
• availability of effective treatment.

These classifications presume ordinary circumstances in the research laboratory or growth in small volumes for diagnostic and experimental purposes. Four levels of risk have been defined as follows⁽¹⁾.

Risk Group 1 (low individual and community risk)

Any biological agent that is unlikely to cause disease in healthy workers or animals.

Risk Group 2 (moderate individual risk, low community risk)

Any pathogen that can cause human disease but, under normal circumstances, is unlikely to be a serious hazard to laboratory workers, the community, livestock or the environment. Laboratory exposures rarely cause infection leading to serious disease; effective treatment and preventive measures are available, and the risk of spread is limited.

Risk Group 3 (high individual risk, low community risk)

Any pathogen that usually causes serious human disease or can result in serious economic consequences but does not ordinarily spread by casual contact from one individual to another, or that causes diseases treatable by antimicrobial or antiparasitic agents.

Risk Group 4 (high individual risk, high community risk)

Any pathogen that usually produces very serious human disease, often untreatable, and may be readily transmitted from one individual to another, or from animal to human or vice-versa, directly or indirectly, or by casual contact.

A list of human pathogens categorized according to Risk Group can be obtained by calling the Office of Laboratory Security directly at (613) 957-1779 or accessing their Web site: http://www.phac-aspc.gc.ca/ols-bsl/

2.2 Containment Levels

Classification of organisms according to risk group is not meant to establish the actual handling of biological hazards in the laboratory setting. For example, the risk group system does not take into account the procedures that are to be employed during the manipulation of a particular organism. Containment levels are selected to provide the end-user with a description of the minimum containment required for handling the organism safely in a laboratory setting. In addition to the inherent characteristics of each organism as described in section 2.1, the containment system includes the engineering, operational, technical and physical requirements for manipulating a particular pathogen⁽²⁾. These containment levels are applicable to facilities such as diagnostic, research, clinical, teaching and production facilities that are working at a laboratory scale (for large scale see Chapter 6). Four containment levels are described as follows:

Containment Level 1 (CL1)

This applies to the basic laboratory that handles agents requiring containment level 1. CL1 requires no special design features beyond those suitable for a well-designed and functional laboratory. Biological safety cabinets (BSCs) are not required. Work may be done on an open bench top, and containment is achieved through the use of practices normally employed in a basic microbiology laboratory.

Containment Level 2 (CL2)

This applies to the laboratory that handles agents requiring containment level 2. The primary exposure hazards associated with organisms requiring CL2 are through the ingestion, inoculation and mucous membrane route. Agents requiring CL2 facilities are not generally transmitted by airborne routes, but care must be taken to avoid the generation of aerosols (aerosols can settle on bench tops and become an ingestion hazard through contamination of the hands⁽³⁾ ) or splashes. Primary containment devices such as BSCs and centrifuges with sealed rotors or safety cups are to be used as well as appropriate personal protective equipment (i.e., gloves, laboratory coats, protective eyewear). As well, environmental contamination must be minimized by the use of handwashing sinks and decontamination facilities (autoclaves).

Containment Level 3 (CL3)

This applies to the laboratory that handles agents requiring containment level 3. These agents may be transmitted by the airborne route, often have a low infectious dose to produce effects and can cause serious or life-threatening disease. CL3 emphasizes additional primary and secondary barriers to minimize the release of infectious organisms into the immediate laboratory and the environment. Additional features to prevent transmission of CL3 organisms are appropriate respiratory protection, HEPA filtration of exhausted laboratory air and strictly controlled laboratory access.

Containment Level 4 (CL4)

This is the maximum containment available and is suitable for facilities manipulating agents requiring containment level 4. These agents have the potential for aerosol transmission, often have a low infectious dose and produce very serious and often fatal disease; there is generally no treatment or vaccine available. This level of containment represents an isolated unit, functionally and, when necessary, structurally independent of other areas. CL4 emphasizes maximum containment of the infectious agent by complete sealing of the facility perimeter with confirmation by pressure decay testing; isolation of the researcher from the pathogen by his or her containment in a positive pressure suit or containment of the pathogen in a Class III BSC line; and decontamination of air and other effluents produced in the facility.

2.3 Risk Assessment

Risk assessment is a critical step in the selection of an appropriate containment level for the microbiological work to be carried out. A detailed local risk assessment should be conducted to determine whether work requires containment level 1, 2, 3 or 4 facilities and operational practices. Individuals with varying expertise and responsibilities should be included in the risk assessment process and can include, among others, the facility director, laboratory supervisor, principal investigator, senior microbiologist, biosafety officer and biosafety committee.

Available information can be used as a starting point to assist in the identification of risk factors, including the recommended Risk Group of the organism (see section 2.1 Risk Groups). In addition to the Risk Group classifications, which are based on the risk factors inherent to the organism, the following factors associated with the laboratory operation should also be examined:

• potential for aerosol generation
• quantity
• concentration
• agent stability in the environment (inherent biological decay rate)
• type of work proposed (e.g., in vitro , in vivo , aerosol challenge studies)
• use of recombinant organisms (e.g., gene coding for virulence factors or toxins; host range alteration; oncogenicity; replication capacity; capability to revert to wild type).

The containment level required for work with a particular agent is based on the manipulations generally associated with laboratory scale research and clinical procedures. If a particular procedure, such as preliminary identification, poses a lower hazard than manipulation of a live culture, then a lower containment level may be appropriate. For example, primary diagnostic tests for HIV may be done in a containment level 2 physical laboratory with the use of containment level 3 operational protocols, but growing and manipulating a culture of HIV may require both containment level 3 physical facility and operational protocols.

On the other hand, an increase in containment may be required if the local risk assessment indicates that the procedures pose a higher risk than routine laboratory scale and diagnostic manipulations. For example, Corynebacterium diphtheriae (aerosol transmitted) may be manipulated for diagnostic work and laboratory scale research in a containment level 2 laboratory; however, animal aerosol inhalation challenges may require increased levels of physical and operational containment.

An increase in containment may be required once a facility begins large scale production. "Large scale" generally refers to volumes manipulated in a single volume in excess of 10 L. Because of the significant quantity of infectious material being handled, special precautions relating specifically to large scale quantities have been developed and are detailed in Chapter 6. It must be noted that the 10 L cut-off is not an absolute value. A hazard analysis may indicate that, because of high pathogenicity, the route of transmission and the low infectious dose, a particular study involving volumes of < 10 L but larger than research scale volumes may pose a greater hazard than research scale quantities and therefore may require increased levels of physical and operational containment. For example, a hazard analysis may indicate that a procedure involving production of 5 L quantities of MDRTB (multi-drug resistant Mycobacterium tuberculosis ) is more appropriately carried out at containment level 3 large scale than at diagnostic and laboratory scale containment level 3. Therefore the 10 L cut-off between laboratory scale and large scale is to be used as a guide only, and a thorough risk assessment should be carried out on a case-by-case basis.

Further guidance on carrying out a risk assessment and related information that can be used to assist in the risk assessment procedure can be found in the Centers for Disease Control and Prevention/National Institutes of Health Biosafety in Microbiological and Biomedical Laboratories⁽⁴⁾. This information is also available by accessing the following Web site: http://www.cdc.gov/od/ohs/

2.4 Health and Medical Surveillance

A health and medical surveillance program (including pre-employment and then periodic testing) needs to be appropriate to the agents in use and the programs in place in the laboratory. As such, the details of the health and medical surveillance program would be determined and defined by a risk assessment process based upon Canadian and International practices^(1,4,5) that clearly demonstrates the reasons, indications and advantages for such a program to be in place. This program may include but is not limited to the following: a medical examination; serum screening, testing and/or storage; immunizations; and possibly other tests as determined by the risk assessment process. Risk assessment should be carried out by a multidisciplinary group including management, safety and occupational health professionals. The health and medical surveillance program risk assessment would include consideration of those people working with high risk organisms, because knowledge of immune status is critical for decisions concerning immunizations, prophylaxis, etc.⁽¹⁾.

Only people meeting these identified medical entry requirements (e.g., immunizations) may enter the laboratory unless the facility has been appropriately decontaminated. Or, other specified protocols can be developed and implemented to achieve the same level of protection for other individuals entering a facility.

2.5 Management of Biological Safety

Although the responsibility for the safety of staff lies with the supervisors and directors of the microbiology laboratory, it can be advantageous to identify an individual(s) to specifically manage biological safety issues. In many laboratories, this role is either informally assigned to a qualified individual who performs these duties on a part-time basis (e.g., senior microbiologist) or the role is shared by a number of individuals. This role can also be formally assigned to a dedicated Biological Safety Officer who has a working knowledge of the laboratory practices and procedures within the facility.

The formation of an Institutional Biosafety Committee to oversee the facility's biological safety program can also be incorporated into the structure for the management of biological safety issues (in some institutions and universities, the requirement for an Institutional Biosafety Committee is mandatory). The Biological Safety Officer (or individual assigned to manage biological safety issues) should liaise with the Committee through regularly scheduled meetings and can present specific safety problems, concerns or policy/protocol improvements to be considered and addressed. The Committee is also available to the Biological Safety Officer for risk assessments, disputes about biological safety matters or other matters that may be of a biological safety nature. Careful consideration is to be given to the composition of the Institutional Biosafety Committee, which, when possible, should include several individuals with varying expertise, the Biological Safety Officer, at least one member of each of the research staff (researcher) and technical staff (technician), and a representative from management. Consideration should also be given to representation by a medical advisor.

The structure for the management of biological safety issues within each facility should be determined locally and will vary according to the level of coordination and the associated resources necessary for implementation. Determining factors include the following:

• the size of the facility (staff and square footage)
• the concentration of multiple laboratories in the facility
• the containment levels within the facility (level 2 laboratory, multiple level 3 laboratories)
• the complexity of the processes (routine diagnostics, research, large scale, recombinant work)
• the existence of shared laboratory space within the facility (multiple investigators, various organizations)
• experimental or diagnostic animal activities within the facility (mice in containment caging, large animal housing).

Biological safety issues to be managed may include the following:

• identifying training needs and assisting with the development and delivery of biosafety training programs, such as general biosafety, BSC use, animal biosafety, staff orientation and containment suite training.
• performing risk assessments when required and developing recommendations for procedural or physical laboratory modifications.
• auditing the effectiveness of the biosafety program and its associated management system on a regular basis.
• participating in accident investigations and promoting the reporting of incidents within the facility or laboratory.
• distributing new and relevant biosafety information to laboratory staff.
• coordinating and monitoring the decontamination, disinfection and disposal procedures for infectious materials in the facility or laboratory.
• coordinating the receipt, shipment and transport within the facility of infectious material according to the Workplace Hazardous Materials Information System (WHMIS) and Transportation of Dangerous Goods (TDG) regulations.
• establishing a record keeping and secure storage system for all infectious material entering the facility.
• coordinating emergency response activities.
• maintaining liaison with support staff, housekeeping staff and contractors on matters related to facility biosafety.

CL 3 or 4 laboratories may have the additional biosafety activities:

• certification and recertification of the laboratory (see Chapter 5).
• investigation and remediation of containment suite physical or operational failures.
• access control to the containment suites.
• liaison with applicable regulatory bodies, such as the Nuclear Safety Commission, Transport Canada, Health Canada and the Canadian Food Inspection Agency.

2.6 Biosecurity

Today, facilities handling infectious agents need not only a biosafety program but also a biosecurity plan in place. While biosafety deals with all aspects of containment to prevent any exposure to and accidental release of pathogens, biosecurity is implemented to prevent the theft, misuse or intentional release of pathogens. Whether it be for the advancement of science or the diagnosis of agents causing disease or the misuse of these technologies, there is unfortunately a dual use potential in the nature of the work (i.e., procedures, equipment, etc.) that takes place with these agents⁽⁶⁾.There are many international recommendations^(7-10) and position papers (11-16) which can provide further assistance with the management of biological threats.

As the planning and implementation of a biosecurity plan needs to be specific to the nature of each facility, the type of research and diagnostics conducted, and the local environment, a diverse working group needs to be involved. Consideration should be made to include scientific directors, principal investigators, laboratory workers, administrators, safety officers, security staff, maintenance staff, and law enforcement agencies where appropriate. Also, include the "Responsible Official" (RO) where one is designated. A Responsible Official is typically responsible for the development, training, and implementation of safety, security, and emergency response plans. As such, the RO is contacted with timely notice of any theft, loss or release of agents. This individual is involved in allowing only approved individuals to have access to agents and is involved in the transfer and transportation of agents from the facility. This person can assist with maintaining detailed records of information necessary to give a complete accounting of all activities related to pathogens.

A primary component to a biosecurity plan must be a detailed risk assessment (see also Chapter 2.3)^(7,10). The biosecurity risk assessment should review and list the relevant assets, define the threats, outline the vulnerabilities, and determine the countermeasures or mitigation strategies specific for each facility. The biosecurity plan should then address the following factors^(8,11,15) : physical protection; personnel suitability/reliability; pathogen accountability; and related incident and emergency response.

Given the importance and timely nature of the issue of biosecurity, this section was added after the final Contributors meeting. However, the specific requirements have been included in the matrices as reviewed by the Contributors.

Physical Protection

The physical protection risk assessment should include all levels of biosecurity review: perimeter security, facility security, laboratory security and agent specific security, and outline procedures for securing the area, e.g., card access, key pads, locks etc. All laboratories should adopt biosecurity practices to minimize opportunities for unauthorized entry into laboratories, animal and storage areas, as well as the unauthorized removal of infectious materials from their facility. Similarly, information security for data and electronic technology need to be addressed.

Personnel Suitability/Reliability

Background checks and security clearances may be required before employees are granted access to containment facilities. These factors should be considered as part of the local risk assessment process when developing a biosecurity plan. Photo identification badges for employees and temporary badges for escorted visitors can also be used to identify individuals with clearance to enter restricted areas. Procedures are needed for approving and granting visitors access to controlled areas. In this capacity the access to agents and storage facilities is limited to legitimate use/individuals only. Biosecurity training needs to be provided to all personnel who are given access.

Pathogen Accountability

Pathogen accountability procedures should include inventory requirements for proper labelling, tracking of internal possession, inactivation and disposal of cultures after use, and transfers within and outside the facility. These inventory controls also assist in keeping track of pathogen storage locations and under whose responsibility the pathogens lie. Inventories must be updated regularly to include new additions as a result of diagnosis, verification of proficiency testing, or receipt from other locations as well as to remove agents after transfers or appropriate inactivation and disposal mechanisms have been used. The record keeping should include pathogen inventories, who has access to agents, who has access to areas where agents are stored or used, as well as transfer documents. A notification process for identifying, reporting, and remediating security problems, i.e., inventory discrepancy, equipment failure, breach of security, release of agents, etc., should be in place.

Biosecurity Incident and Emergency Response

A protocol for reporting and investigating security incidents e.g., missing infectious substances, unauthorized entry, should be addressed. A mechanism needs to be in place for the reporting and removal of unauthorized persons. Biosecurity incident and emergency plans should include response to intentional (bomb threats etc.), unintentional (accidental release) and natural events (power outages, severe weather). Training needs to be provided to all relevant personnel.

Biosecurity requirements for facilities handling infectious agents at containment levels 3 and 4 will generally be more stringent than those required in clinical and research containment level 2 laboratories. Recommendations on biosecurity practices (e.g., storage of pathogens, inventories, log books to record entry) and physical design security features (locks, restricted access) have been incorporated into the requirements for each containment level in Chapters 3 and 4.

Expert advice from security and/or law enforcement experts should be sought in the development of threat assessments and security protocols specific to each facility. The threat assessment and security practices should be regularly reviewed and updated to reflect new threats that may be identified.

References

1. Laboratory biosafety manual. Geneva: World Health Organization, 1993;1-133.

2. Collins, C.H., and Kennedy, D.A. Equipment-and technique-related hazards. In: Laboratory-acquired infections: history, incidence, causes and preventions. Oxford, U.K: Butterworth-Heinemann, 1999; 65-109.

3. Collins, C.H., and Kennedy, D.A. Exposure, sources and routes of infection. In: Laboratory-acquired infections: history, incidence, causes and preventions. Oxford, U.K: Butterworth-Heinemann, 1999; 38-53.

4. Richmond, J.Y. and McKinney, R.W. Biosafety in microbiological and biomedical laboratories. Washington, DC: U.S. Government Printing Office, 1999;1-250.

5. The management, design and operation of microbiological containment laboratories. Advisory Committee on Dangerous Pathogens, UK: Health and Safety Executive, 2001; 7-16.

6. Biotechnology Research in an Age of Terrorism : Confronting the Dual Use Dilemma. Washington, DC: The National Academic Press, 2004.

7. Public Health Response to Biological and Chemical Weapons: WHO Guidance. Geneva: World Health Organization, Draft 2003.

8. Interim Final Rule: CDC; Possession, Use, and Transfer of Select Agents and Toxins. Federal Register, Department of Health and Human Services, 2002; 240 (67): 76885-76905.

9. Richmond, J.Y. and McKinney, R.W. Biosafety in microbiological and biomedical laboratories. Washington, DC: U.S. Government Printing Office, 1999;1-250.

10. Laboratory Security and Emergency Response Guidance for Laboratories Working with Select Agents. MMWR, 2002; 51(RR-19):1-6.

11. ABSA Biosecurity Task Force White Paper: Understanding Biosecurity. American Biological Safety Association, January   2003. (http://www.absa.org/0301bstf.html).

12. Biosafety, Biosecurity, and Biological Weapons. Germany: The Sunshine Project, October 2003;1-22.

13. Salerno, R., and Koelm, J.G. Biological Laboratory and Transportation Security and the Biological Weapons Convention. Albuquerque, New Mexico: Sandia National Laboratories, SAND No.2002-1067P, February 2002; 1-9. (http://cns.miis.edu/ research/cbw/biosec/pdfs/sandia.pdf).

14. Barletta, M., Biosecurity Measures for Preventing Bioterrorism. Center for Nonproliferation Studies, Monterey Institute of International Studies, 2002; 1-11.

15. Tucker, J.B., Biosecurity: Limiting Terrorist Access to Deadly Pathogens. Washington, DC: United States Institute of Peace, Peaceworks, 2003; 1-51.

16. Controls Over Biological, Chemical, and Radioactive Materials at Institutions Funded by the U.S. Department of Agriculture. Washington, DC: United States Department of Agriculture, Report No. 50099-14-At, September 2003; 1-43. (http://www.usda.gov/oig/webdocs/50099-14-At.pdf).

[Previous] [Table of Contents] [Next]