Biosafety Manual
2015
Policy Statement
It is the policy of Northern Arizona University (NAU) to provide a safe working environment. The
primary responsibility for insuring safe conduct and conditions in the laboratory resides with the
principal investigator.
The Biological Safety Office is committed to providing up-to-date information, training, and
monitoring to the research and biomedical community concerning the safe conduct of biological,
recombinant, and acute toxin research and the handling of biological materials in accordance with all
pertinent local, state and federal regulations, guidelines, and laws. To that end, this manual is a
resource, to be used in conjunction with the CDC and NIH guidelines, the NAU Select Agent
Program, Biosafety in Microbiological and Biomedical Laboratories (BMBL), and other resource
materials.
Introduction
This Biological Safety Manual is intended for use as a guidance document for researchers and
clinicians who work with biological materials. It should be used in conjunction with the LaboratorySpecific Safety Manual, which provides more general safety information. These manuals describe
policies and procedures that are required for the safe conduct of research at NAU. The NAU
Personnel Policy on Safety 5.03 also provides guidance for safety in the workplace.
Responsibilities
In the academic research/teaching setting, the principal investigator (PI) is responsible for ensuring
that all members of the laboratory are familiar with safe research practices. In the clinical laboratory
setting, the faculty member who supervises the laboratory is responsible for safety practices.
Lab managers, supervisors, technicians and others who provide supervisory roles in laboratories
and clinical settings are responsible for overseeing the safety practices in laboratories and reporting
any problems, accidents, and spills to the appropriate faculty member.
Employees who work with biological materials are responsible for reading this manual and carrying
out the safety practices outlined here.
The Biosafety Office will provide guidance, information, review, monitoring, and training regarding
biological safety programs, when appropriate. This includes implementing registration activities for
certain research projects, acting as a consultant for departments regarding implementation and
enforcement of biological safety programs, evaluating work practices and personal protective
equipment, providing educational materials, tracking employee training, and medical monitoring.
The mission of the EH&S Biosafety Office is to minimize injury to faculty, staff, students and visitors
and to minimize damage to University property. Inherent in this mission is the charge to provide a
safe and healthy environment in which the University’s activities can be pursued.
All applicable federal and state safety laws, rules and regulations are adopted by the University. In
order to carry out its duties and responsibilities, the EH&S Biosafety Office will adhere to standards
or codes related to biosafety which have been adopted and promulgated by nationally recognized
standards-setting organizations. The interpretation of biosafety regulations and guidelines is the
responsibility of the EH&S Biosafety Office.
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In order to assure an effective Biosafety Program for the Northern Arizona University, it is imperative
that all individuals associated with the University comply fully with the policies and procedures set
forth in the manual.
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Emergency Phone Numbers
General
University Police Department
523-3000
Flagstaff Fire Department
911
Gas leak
911
Spills/Accidents
Biological or Recombinant Materials
480-248-0741
Chemicals (laboratory)
928-607-9995
Radioactive Materials
928-220-1388
Select Agents
480-248-0741
Incidents involving Air Monitoring
928-220-1728
Medical Emergency
Campus Health Services
523-2131
Flagstaff Medical Center Emergency
779-3366
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Table of Contents
1 – Biological Safety ................................................................................................................10
Principles of Biological Safety ...........................................................................................10
Laboratory Practice and Technique...................................................................................10
Safety Equipment (Primary Barrier) .................................................................................11
Facility Design (Secondary Barrier)..................................................................................11
Biosafety Levels ................................................................................................................12
Animal Facilities ...............................................................................................................14
Clinical Laboratories .........................................................................................................14
Importation and Interstate Shipment of Certain Biomedical Materials ............................15
Biological Safety Levels ......................................................................................................15
Biosafety Level 1 ..............................................................................................................15
Biosafety Level 2 ..............................................................................................................17
Biosafety Level 3 ..............................................................................................................20
Animal Biosafety Level 1 (ABSL-1) ................................................................................25
Animal Biosafety Level 2 (ABSL-2) ................................................................................27
Animal Biosafety Level 3 (ABSL-3) ................................................................................30
Agents List ...........................................................................................................................37
Biological Safety Level 1 (BSL-1)....................................................................................37
Biological Safety Level 2 (BSL-2)....................................................................................37
BSL-2 - Bacterial Agents Including Chlamydia ....................................................... 38
BSL-2 - Fungal Agents ............................................................................................. 39
BSL-2 - Parasitic Agents........................................................................................... 39
BSL-2 - Viruses ........................................................................................................ 40
Biological Safety Level 3 (BSL-3)....................................................................................42
BSL-3 - Bacterial Agents Including Rickettsia ......................................................... 42
BSL-3 - Fungal Agents ............................................................................................. 42
BSL-3 - Parasitic Agents........................................................................................... 42
BSL-3 - Viruses and Prions ...................................................................................... 43
Biological Safety Level 4 (BSL-4)....................................................................................44
BSL-4 - Bacterial Agents .......................................................................................... 44
BSL-4 - Fungal Agents ............................................................................................. 44
BSL-4 - Parasitic Agents........................................................................................... 44
BSL-4 - Viral Agents ................................................................................................ 44
2 - Information for Researchers ..............................................................................................45
Project Registration ............................................................................................................45
Biological Agent (BA) Registration..................................................................................45
Recombinant DNA (R-DNA) Registration .......................................................................45
Acute Toxins (AT) Registration .......................................................................................46
Regulated Biological Materials .........................................................................................46
Project Amendment ..........................................................................................................46
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Typical changes include: ...................................................................................................46
Table 3: Toxin Table .........................................................................................................47
Select Agents ....................................................................................................................48
Minors in Research Laboratories or Animal Facilities ...................................................50
Responsibilities of the Principle Investigator during the Conduct of the Research .....51
Reportable Incidents and Violations .................................................................................52
Institutional Reporting Responsibilities ............................................................................52
Biological Waste Disposal Policy ............................................................................................53
Categories ............................................................................................................................53
1) Infectious/potentially infectious/R-DNA .................................................................. 53
2) Non-infectious waste ................................................................................................ 53
3) Mixed radioactive/biohazardous waste ................................................................ 53
4) Mixed chemical/biohazardous waste .................................................................... 54
5) Animal carcasses and materials ............................................................................ 54
6) Human remains ..................................................................................................... 54
Packaging ..........................................................................................................................54
1) Biohazard bags...................................................................................................... 54
2) Sharps ................................................................................................................... 54
3) Containers ........................................................................................................... 54
Labeling ............................................................................................................................54
1) Date ....................................................................................................................... 54
2) Name/Location/Phone Number ............................................................................ 55
3) Biohazard sign ...................................................................................................... 55
Transport ...........................................................................................................................55
Training .............................................................................................................................55
Biological Waste Disposal Containers ....................................................................................55
Bulk Biohazard Receptacles .............................................................................................56
Sharps Boxes .....................................................................................................................56
Biohazard Bags .................................................................................................................56
Autoclave Use and Testing ......................................................................................................57
Autoclave Testing .............................................................................................................57
Record-Keeping ................................................................................................................58
Autoclave Operating Procedures ......................................................................................58
Autoclave Operation and Safety Training.........................................................................59
Autoclave Guidelines ........................................................................................................60
Disinfectants .............................................................................................................................61
Liquids ..............................................................................................................................61
Alcohols .................................................................................................................... 61
Chlorine compounds ................................................................................................. 62
Formaldehyde............................................................................................................ 62
Glutaraldehyde .......................................................................................................... 62
Hydrogen peroxide .................................................................................................... 62
Iodine and Iodophors................................................................................................. 62
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Phenol and phenolic compounds .............................................................................. 63
Quatenary ammonium compounds ........................................................................... 63
Gases .................................................................................................................................63
Ethylene Oxide (Not currently allowed at NAU)...................................................... 63
Vapor Phase Hydrogen Peroxide .............................................................................. 63
Chlorine Dioxide gas ................................................................................................ 63
Ozone ........................................................................................................................ 63
Formaldehyde Gas (from heating paraformaldehyde) .............................................. 64
Irradiation ..........................................................................................................................64
Ultraviolet, UV radiation .......................................................................................... 64
Ionizing radiation ...................................................................................................... 64
Electron Beam ........................................................................................................... 64
Microwaves ............................................................................................................... 64
Disinfectants Bibliography................................................................................................64
Shipment of Biological Materials ............................................................................................69
Laboratory Spills ......................................................................................................................70
Spill in the Open Laboratory .............................................................................................70
Spill in a Biological Safety Cabinet ..................................................................................71
Biological Safety Cabinets .......................................................................................................72
Biological Safety Cabinets (BSCs) ...................................................................................72
The Class I BSC ........................................................................................................ 72
The Class II BSC....................................................................................................... 72
The Class II, Type A BSC ......................................................................................... 73
The Class II, Type B1 BSC ....................................................................................... 73
The Class II, Type B2 BSC ....................................................................................... 73
The Class II, Type B3 BSC ....................................................................................... 74
The Class III BSC ..................................................................................................... 74
Horizontal Laminar Flow “Clean Bench” .........................................................................74
Vertical Laminar Flow “Clean Bench” .............................................................................74
Operations within a Class II BSC .....................................................................................75
Laboratory Hazards ................................................................................................... 75
Decontamination ...............................................................................................................76
Surface Decontamination .......................................................................................... 76
Gas Decontamination ................................................................................................ 77
Engineering Requirements ................................................................................................77
Ultraviolet Lamps ..................................................................................................... 77
BSC Placement ......................................................................................................... 77
HEPA Filters ............................................................................................................. 77
Certification of BSCs ................................................................................................ 77
3 – Programs ............................................................................................................................78
Animal Contact Medical Monitoring Program ......................................................................78
NAU Bloodborne Pathogen Program ................................................................................80
BBP Exposure Information ....................................................................................... 80
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4 -Medical Surveillance ...........................................................................................................81
HIV Research Laboratory Occupational Medicine Policy .............................................82
Pre-employment ................................................................................................................82
Continuing employment ....................................................................................................82
Post-exposure prophylaxis ................................................................................................82
References .........................................................................................................................83
Immunoprophylaxis ............................................................................................................84
Recommendations for Prophylactic Immunization of Laboratory Personnel Working with
Infectious Agents ..............................................................................................................85
Bacterial agents ......................................................................................................... 85
Rickettsial agents ...................................................................................................... 86
Viral agents ............................................................................................................... 86
Vaccinia Immunization Policy ...........................................................................................87
Blood Serum Sampling .....................................................................................................88
Health Assessments ..........................................................................................................88
Exposure to Mycobacterium Tuberculosis .......................................................................88
5 – Appendices - Information for Researchers .......................................................................89
Transgenic Animals ............................................................................................................89
List of Tables
Table 1: Summary of Recommended Biosafety Levels for Infectious Agents ......................... 35
Table 2: Summary of Recommended Biosafety Levels for Activities in Which
Experimentally or Naturally Infected Vertebrate Animals Are Used ............................... 36
Table 3: Toxin Table................................................................................................................. 47
Table 4: Dilutions of Household Bleach ................................................................................... 62
Table 5: Summary and Comparison of Liquid Disinfectants (Page 1) ..................................... 65
Table 6: Summary of Practical Disinfectants............................................................................ 67
Table 7: Reprocessing Methods for Equipment Used in the Health Care Setting .................... 68
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1 – Biological Safety
Principles of Biological Safety
The following is from Biosafety in Microbiological and Biomedical Laboratories, 2009, HHS publication No. (CDC) 21-1112,
Centers for Disease Control & Prevention/National Institutes of Health
The term “containment” is used in describing safe methods for managing infectious agents in the
laboratory environment where they are being handled or maintained. The purpose of
containment is to reduce or eliminate exposure of laboratory workers, other persons, and the
outside environment to potentially hazardous agents.
Both good microbiological technique and the use of appropriate safety equipment provide
primary containment, the protection of personnel and the immediate laboratory environment from
exposure to infectious agents. The use of vaccines may provide an increased level of personal
protection. Secondary containment, the protection of the environment external to the laboratory
from exposure to infectious materials, is provided by a combination of facility design and
operational practices. Therefore, the three elements of containment include laboratory practice
and technique, safety equipment, and facility design. The risk assessment of the work to be done
with a specific agent will determine the appropriate combination of these elements.
Laboratory Practice and Technique
The most important element of containment is strict adherence to standard microbiological
practices and techniques. Persons working with infectious agents or potentially infectious
materials must be aware of potential hazards, and must be trained and proficient in the practices
and techniques required for handling such material safely. The director or person in charge of the
laboratory is responsible for providing or arranging for appropriate training of personnel.
Each laboratory should develop or adopt a biosafety or operations manual that identifies the
hazard that will or may be encountered, and which specifies practices and procedures designed to
minimize or eliminate risks. Personnel should be advised of special hazards and should be
required to read and to follow the required practices and procedures. A scientist trained and
knowledgeable in appropriate laboratory techniques, safety procedures, and hazards associated
with handling infectious agents must direct laboratory activities.
When standard laboratory practices are not sufficient to control the hazard associated with a
particular agent or laboratory procedure, additional measures may be needed. The laboratory
director is responsible for selecting additional safety practices, which must be in keeping with the
hazard associated with the agent or procedure.
Laboratory personnel, safety practices, and techniques must be supplemented by appropriate
facility design and engineering features, safety equipment, and management practices.
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Safety Equipment (Primary Barrier)
Safety equipment includes biological safety cabinets (BSCs), enclosed containers, and other
engineering controls designed to remove or minimize exposures to hazardous biological
materials. The biological safety cabinet (BSC) is the principal device used to provide
containment of infectious splashes or aerosols generated by many microbiological procedures.
Three types of BSCs (Class I, II, III) used in microbiological laboratories are described in Section
2. Open-fronted Class I and Class II BSCs are primary barriers which offer significant levels of
protection to laboratory personnel and to the environment when used with good microbiological
techniques. The Class II BSC also provides protection from external contamination of the
materials (e.g., cell cultures, microbiological stocks) being manipulated inside the cabinet. The
gas-tight Class III BSC provides the highest attainable level of protection to personnel and the
environment.
An example of another primary barrier is the safety centrifuge cup, an enclosed container
designed to prevent aerosols from being released during centrifugation. To minimize this hazard,
containment controls such as BSC's or centrifuge cups must be used for handling infectious
agents that can be transmitted through the aerosol route of exposure.
Safety equipment also may include items for personal protection such as gloves, coats, gowns,
shoe covers, boots, respirators, face shields, safety glasses, or goggles. Personal protective
equipment is often used in combination with biological safety cabinets and other devices that
contain the agents, animals, or materials being worked with. In some situations in which it is
impractical to work in biological safety cabinets, personal protective equipment may form the
primary barrier between personnel and the infectious materials. Examples include certain animal
studies, animal necropsy, agent production activities, and activities relating to maintenance,
service, or support of the laboratory facility.
Facility Design (Secondary Barrier)
The design of the facility is important in providing a barrier to protect persons working inside
and outside of the laboratory within the facility, and to protect persons or animals in the
community from infectious agents that may be accidentally released from the laboratory.
Laboratory management is responsible for providing facilities commensurate with the
laboratory's function and the recommended biosafety level for the agents being manipulated.
The recommended secondary barrier(s) will depend on the risk of transmission of specific agents.
For example, the exposure risks for most laboratory work in Biosafety Level 1 and 2 facilities
will be direct contact with the agents, or inadvertent contact exposures through contaminated
work environments. Secondary barriers in these laboratories may include separation of the
laboratory work area from public access, availability of a decontamination facility (e.g.,
autoclave), and hand washing facilities.
As the risk for aerosol transmission increases, higher levels of primary containment and multiple
secondary barriers may become necessary to prevent infectious agents from escaping into the
environment. Such design features could include specialized ventilation systems to assure
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directional air flow, air treatment systems to decontaminate or remove agents from exhaust air,
controlled access zones, airlocks as laboratory entrances, or separate buildings or modules for
isolation of the laboratory. Design engineers for laboratories may refer to specific ventilation
recommendations as found in the Applications Handbook for Heating, Ventilation, and Airconditioning (HVAC) published by the American Society of Heating, Refrigerating, and Airconditioning Engineers (ASHRAE).
Biosafety Levels
Four biosafety levels (BSLs) are described which consist of combinations of laboratory practices
and techniques, safety equipment, and laboratory facilities. Each combination is specifically
appropriate for the operations performed, the documented or suspected routes of transmission of
the infectious agents, and for the laboratory function or activity.
The recommended BSLs for the organisms represent those conditions under which the agent can
ordinarily be safely handled. The Principle Investigator (PI) is specifically and primarily
responsible for assessing risks and for appropriately applying the recommended biosafety levels.
Generally, work with known agents should be conducted at the biosafety level recommended.
When specific information is available to suggest that virulence, pathogenicity, antibiotic
resistance patterns, vaccine and treatment availability, or other factors are significantly altered,
more (or less) stringent practices may be specified.
Biosafety Level 1 practices, safety equipment, and facilities are appropriate for undergraduate
and secondary educational training and teaching laboratories, and for other facilities in which
work is done with defined and characterized strains of viable microorganisms not known to cause
disease in healthy adult humans. Bacillus subtilis, Naegleria gruberi, and infectious canine
hepatitis virus are representative of those microorganisms meeting these criteria. Many agents
not ordinarily associated with disease processes in humans are, however, opportunistic pathogens
and may cause infection in the young, the aged, and immunodeficient or immunosuppressed
individuals. Vaccine strains which have undergone multiple in vivo passages should not be
considered avirulent simply because they are vaccine strains.
Biosafety Level 1 represents a basic level of containment that relies on standard microbiological
practices with no special primary or secondary barriers recommended, other than a sink for hand
washing.
Biosafety Level 2 practices, equipment, and facilities are applicable to clinical, diagnostic,
teaching, and other facilities in which work is done with the broad spectrum of indigenous
moderate-risk agents present in the community and associated with human disease of varying
severity. With good microbiological techniques, these agents can be used safely in activities
conducted on the open bench, provided the potential for producing splashes or aerosols is low.
Salmonellae, clinical samples of Hepatitis B, and Toxoplasma spp. are representative of
microorganisms assigned to this containment level. Biosafety Level 2 is appropriate when work
is done with any human-derived blood, body fluids, or tissues where the presence of an infectious
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agent may be unknown. (Laboratory personnel working with human-derived materials should
refer to the Bloodborne Pathogen Standard for specific, required precautions).
Primary hazards to personnel working with these agents relate to
accidental percutaneous or mucous membrane exposures, or ingestion of
infectious materials. Extreme precaution with contaminated needles or
sharp instruments must be emphasized. Even though organisms routinely
manipulated at BSL-2 are not known to be transmissible by the aerosol
route, procedures with aerosol or high splash potential that may increase
the risk of such personnel exposure must be conducted in primary containment equipment, or
devices such as a BSC or safety centrifuge cups. Other primary barriers should be used, as
appropriate, such as splash shields, face protection, gowns, and gloves.
Secondary barriers such as hand washing and waste decontamination facilities must be available
to reduce potential environmental contamination.
Biosafety Level 3 practices, safety equipment, and facilities are applicable to clinical, diagnostic,
teaching, research, or production facilities in which work is done with indigenous or exotic
agents with a potential for respiratory transmission, and which may cause serious and potentially
lethal infection. Hepatitis B. (propagation of high concentrations only), Mycobacterium
tuberculosis, St. Louis encephalitis virus, and Coxiella burnetii are representative of
microorganisms assigned to this level. Primary hazards to personnel working with these agents
relate to autoinoculation, ingestion, and exposure to infectious aerosols.
At Biosafety Level 3, more emphasis is placed on primary and secondary barriers to protect
personnel in contiguous areas, the community, and the environment from exposure to potentially
infectious aerosols. For example, all laboratory manipulations should be performed in a BSC or
other enclosed equipment, such as a gas-tight aerosol generation chamber. Secondary barriers for
this level include controlled access to the laboratory and a specialized ventilation system that
minimizes the release of infectious aerosols from the laboratory.
Biosafety Level 4 practices, safety equipment, and facilities are applicable for work with
dangerous and exotic agents that pose a high individual risk of life-threatening disease, which
may be transmitted via the aerosol route, and for which there is no available vaccine or therapy.
Additionally, agents with a close or identical antigenic relationship to Biosafety Level 4 agents
should also be handled at this level. When sufficient data are obtained, work with these agents
may continue at this level or at a lower level. Viruses
such as Marburg or Congo-Crimean hemorrhagic fever are manipulated at Biosafety Level 4.
The primary hazards to personnel working with Biosafety Level 4 agents are respiratory exposure
to infectious aerosols, mucous membrane exposure to infectious droplets, and autoinoculation.
All manipulations of potentially infectious diagnostic materials, isolates, and naturally or
experimentally infected animals pose a high risk of exposure and infection to laboratory
personnel, the community, and the environment.
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The laboratory worker's complete isolation of aerosolized infectious materials is accomplished
primarily by working in a Class III BSC or a full-body, air-supplied positive-pressure personnel
suit. The Biosafety Level 4 facility itself is generally a separate building or completely isolated
zone with complex, specialized ventilation and waste management systems to prevent release of
viable agents to the environment.
The laboratory director is specifically and primarily responsible for the safe operation of the
laboratory. His/her knowledge and judgment are critical in assessing risks and appropriately
applying these recommendations. The recommended biosafety level represents those conditions
under which the agent can ordinarily be safely handled. Special characteristics of the agents
used, the training and experience of personnel, and the nature or function of the laboratory may
further influence the director in applying these recommendations.
Animal Facilities
Four biosafety levels are also described for activities involving infectious disease work with
experimental mammals. These four combinations of practices, safety equipment, and facilities
are designated Animal Biosafety Levels 1, 2, 3, and 4, and provide increasing levels of protection
to personnel and the environment.
Clinical Laboratories
Clinical laboratories, especially those in health care facilities, receive clinical specimens with
requests for a variety of diagnostic and clinical support services. Typically, the infectious nature
of clinical material is unknown, and specimens are often submitted with a broad request for
microbiological examination for multiple agents (e.g., sputa submitted for “routine,” acid-fast,
and fungal cultures). It is the responsibility of the laboratory director to establish standard
procedures in the laboratory that realistically address the issue of the infectious hazard of clinical
specimens.
Except in extraordinary circumstances (e.g., suspected hemorrhagic fever), the initial processing
of clinical specimens and identification of isolates can be done safely at Biosafety Level 2, the
recommended level for work with bloodborne pathogens such as hepatitis B virus and HIV. The
containment elements described in Biosafety Level 2 are consistent with the Occupational
Exposure to Bloodborne Pathogens Standard from the Occupational Safety and Health
Administration (OSHA), that requires the use of specific precautions with all clinical specimens
of blood or other potentially infectious material (Universal Precautions). Additionally, other
recommendations specific for clinical
laboratories may be obtained from the National Committee for Clinical Laboratory Standards.
Biosafety Level 2 recommendations and OSHA requirements focus on the prevention of
percutaneous and mucous membrane exposures to clinical material. Primary barriers such as
biological safety cabinets (Class I or II) should be used when performing procedures that might
cause splashing, spraying, or splattering of droplets. Biological safety cabinets should also be
used for the initial processing of clinical specimens when the nature of the test requested or other
information is suggestive that an agent readily transmissible by infectious aerosols is likely to be
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present (e.g., M. tuberculosis), or when the use of a biological safety cabinet (Class II) is
indicated to protect the integrity of the specimen.
The segregation of clinical laboratory functions and limiting or restricting access to such areas is
the responsibility of the laboratory director. It is also the director's responsibility to establish
standard, written procedures that address the potential hazards and the required precautions to be
implemented.
Importation and Interstate Shipment of Certain Biomedical Materials
The importation of etiologic agents and vectors of human diseases is subject to the requirements
of the Public Health Service Foreign Quarantine regulations. Companion regulations of the
Public Health Service and the Department of Transportation specify packaging, labeling, and
shipping requirements for etiologic agents and diagnostic specimens shipped in interstate
commerce.
The U. S. Department of Agriculture regulates the importation and interstate shipment of animal
pathogens and prohibits the importation, possession, or use of certain exotic animal disease
agents that pose a serious disease threat to domestic livestock and poultry.
USDA/APHIS permits are required for both importation and within USA transport of many
hazardous biological agents. EH&S maintains all permits and shall be contacted for information
related to any transport to and from NAU. Please see the section entitled “Shipment of
Biological Materials” for more information.
Biological Safety Levels
The following is from Biosafety in Microbiological and Biomedical Laboratories, 2009, HHS publication No. (CDC) 21-1112,
Centers for Disease Control & Prevention/National Institutes of Health
Biosafety Level 1
Biosafety Level 1 is suitable for work involving well-characterized agents not known to cause
disease in healthy adult humans, and of minimal potential hazard to laboratory personnel and the
environment. The laboratory is not necessarily separated from the general traffic patterns in the
building. Work is generally conducted on open bench tops using standard microbiological
practices. Special containment equipment or facility design is not 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 microbiology or a related science.
The following standard and special practices, safety equipment, and facilities apply to the
handling of agents assigned to Biosafety Level 1:
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A.
Standard Microbiological Practices
1.
Access to the laboratory is limited or restricted at the discretion of the laboratory
director when experiments or work with cultures and specimens are in progress.
2.
Persons wash their hands after they handle viable
materials and animals, after removing gloves, and
before leaving the laboratory.
3.
Eating, drinking, use of tobacco products, handling
contact lenses, and applying cosmetics are not
permitted in the work areas where there is
reasonable likelihood of exposure to potentially infectious materials. Persons who
wear contact lenses in laboratories should also wear goggles or a face shield.
Food is stored outside the work area in cabinets or refrigerators designated and
used for that purpose only.
4.
Mouth pipetting is prohibited; mechanical pipetting devices are used.
5.
All procedures are performed carefully to minimize the creation of splashes or
aerosols.
6.
Work surfaces are decontaminated at least once a day and after any spill of viable
material.
7.
All cultures, stocks, and regulated wastes are decontaminated before disposal by
an approved decontamination method, such as autoclaving. Materials to be
decontaminated outside of the immediate laboratory are to be placed in a durable,
leak-proof container and closed for transport from the laboratory. Materials to be
decontaminated off-site from the laboratory are packaged in accordance with
applicable local, state, and federal regulations, before removal from the facility.
8.
An insect and rodent control program is in effect.
B.
Special Practices: None
C.
Safety Equipment (Primary Barriers)
1.
Special containment devices or equipment such as a biological safety cabinet are
generally not required for manipulations of agents assigned to Biosafety Level 1.
2.
It is recommended that laboratory coats, gowns, or uniforms are worn to prevent
contamination or soiling of street clothes.
3.
Gloves should be worn if the skin on the hands is broken or if a rash exists.
4.
Protective eyewear should be worn for anticipated splashes of microorganisms or
other hazardous materials to the face.
D.
Laboratory Facilities (Secondary Barriers)
1.
Each laboratory contains a sink for hand washing.
2.
The laboratory is designed so that it can be easily cleaned. Walls and floors must
be constructed of water impervious materials that will stand up to hard
disinfectants. Carpeting is not allowed in laboratories.
3.
Bench tops are impervious to water and resistant to acids, alkalis, organic
solvents, and moderate heat.
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4.
5.
Laboratory furniture is sturdy and should not be cloth upholstered. Spaces
between benches, cabinets, and equipment are accessible for cleaning.
If the laboratory has windows that open, they are fitted with fly screens.
Biosafety Level 2
Biosafety Level 2 is similar to Level 1 and is suitable for work involving agents of moderate
potential hazard to personnel and the environment. It differs in four ways.
1. Laboratory personnel must have specific training in handling pathogenic agents and
should be directed by competent scientists.
2. Access to the laboratory must be limited when work is being conducted.
3. Extreme precautions must be taken with contaminated sharp items.
4. Certain procedures in which infectious aerosols or splashes may be created must be
conducted in biological safety cabinets or other physical containment equipment.
The following standard and special practices, safety equipment, and facilities apply to the use of
agents assigned to Biosafety Level 2:
A.
Standard Microbiological Practices
1.
Access to the laboratory is limited or restricted at the discretion of the laboratory
director when experiments are in progress.
2.
Persons wash their hands after they handle viable materials and animals, after
removing gloves, and before leaving the laboratory.
3.
Eating, drinking, the use of tobacco products, handling contact lenses, and
applying cosmetics are not permitted in the work areas. Persons who wear contact
lenses in laboratories should also wear goggles or a face shield. Food is stored
outside the work area in cabinets or refrigerators designated for that purpose only.
4.
Mouth pipetting is prohibited; mechanical pipetting devices are used.
5.
All procedures are performed carefully to minimize the creation of splashes or
aerosols.
6.
Work surfaces are decontaminated at least once a day and after any spill of viable
material.
7.
All cultures, stocks, and regulated wastes are decontaminated before disposal by
an approved decontamination method, such as autoclaving. Materials to be
decontaminated outside of the immediate laboratory are to be placed in a durable,
leak-proof container and closed for transport from the laboratory. Materials to be
decontaminated off-site from the laboratory are packaged in accordance with
applicable local, state, and federal regulations, before removal from the facility.
8.
An insect and rodent control program is in effect.
B.
Special Practices
1.
Access to the laboratory is limited or restricted by the laboratory director when
work with infectious agents is in progress. In general, persons who are at
increased risk of acquiring infection or for whom infection may be unusually
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5.
6.
7.
8.
18
hazardous are not allowed in the laboratory or animal rooms. For example,
persons who are immunocompromised or immunosuppressed may be at risk of
acquiring infections. The laboratory director has the final responsibility for
assessing each circumstance and determining who may enter or work in the
laboratory.
2.
The laboratory director establishes policies and procedures whereby only persons
who have been advised of the potential hazard and meet specific entry
requirements (e.g., immunization) enter the laboratory or animal rooms.
3.
When the infectious agent(s) in use in the laboratory require special provisions or
entry (e.g., immunization), a hazard warning sign incorporating the universal
biohazard symbol is posted on the access door to the laboratory work area. The
hazard warning sign identifies the infectious agent, lists the name and telephone
number of the laboratory director or other responsible person(s), and indicates the
special requirement(s) for entering the laboratory.
4.
Laboratory personnel receive appropriate immunizations or tests for the agents
handled or potentially present in the laboratory (e.g., hepatitis B vaccine or TB
skin testing).
When appropriate, considering the agent(s) handled, baseline serum samples for
laboratory and other at-risk personnel are collected and stored. Additional serum
specimens may be collected periodically, depending on the agents handled or the function
of the facility.
A Biosafety Manual is prepared or adopted. Personnel are advised of special hazards and
are required to read and to follow instructions on practices and procedures.
Laboratory personnel receive appropriate training on the potential hazards associated with
the work involved, the necessary precautions to prevent exposures, and the exposure
evaluation procedures. Personnel receive annual updates, or additional training as
necessary for procedural or policy changes.
A high degree of precaution must always be taken with any contaminated sharp items,
including needles and syringes, slides, pipettes, capillary tubes, and scalpels. Needles and
syringes or other sharp instruments should be restricted in the laboratory for use only
when there is no alternative, such as parenteral injection, phlebotomy, or aspiration of
fluids from laboratory animals and diaphragm bottles. Plasticware should be substituted
for glassware whenever possible.
a.
Only needle-locking syringes or disposable syringe-needle units (i.e., needle is
integral to the syringe) are used for injection or aspiration of infectious materials.
Used disposable needles must not be bent, sheared, broken, recapped, removed
from disposable syringes, or otherwise manipulated by hand before disposal;
rather, they must be carefully placed in conveniently located puncture-resistant
containers used for sharps disposal. Non-disposable sharps must be placed in a
hard-walled container for transport to a processing area for decontamination,
preferable by autoclaving.
b.
Syringes that re-sheathe the needle, needle-less systems, and other safe devices
should be used when appropriate.
c.
Broken glassware must not be handled directly by hand, but must be removed by
mechanical means such as a brush and dustpan, tongs, or forceps. Containers of
Biosafety Manual
9.
10.
11.
12.
C.
contaminated needles, sharp equipment, and broken glass are decontaminated
before disposal, according to any local, state, or federal regulations.
Cultures, tissues, or specimens of body fluids are placed in a container that prevents
leakage during collection, handling, processing, storage, transport, or shipping.
Laboratory equipment and work surfaces should be decontaminated with an appropriate
disinfectant on a routine basis, after work with infectious materials is finished, and
especially after overt spills, splashes, or other contamination by infectious materials.
Contaminated equipment must be decontaminated according to any local, state, or federal
regulations before it is sent for repair or maintenance. Equipment must also be
decontaminated before removal from the facility when it must be packaged for transport.
Packaging and shipment shall be in accordance with applicable local, state, or federal
regulations.
Spills and accidents that result in overt exposures to infectious materials are immediately
reported to the laboratory director. Medical evaluation, surveillance, and treatment are
provided as appropriate and written records are maintained.
Animals not involved in work being performed aren’t permitted in the lab.
Safety Equipment (Primary Barriers)
1.
Properly maintained biological safety cabinets, preferably Class II, or other
appropriate personal protective equipment or physical containment devices are
used under the following conditions.
a.
Use Biological Safety Cabinets whenever procedures with a potential for
creating infectious aerosols or splashes are conducted. These may include
centrifuging, grinding, blending, vigorous shaking or mixing, sonic
disruption, opening containers of infectious materials whose internal
pressures may be different from ambient pressures, inoculating animals
intranasally, and harvesting infected tissues from animals or eggs.
b.
Use Biological Safety Cabinets whenever high concentrations or large
volumes of infectious agents are used. Such materials may be centrifuged
in the open laboratory if sealed rotor heads or centrifuge safety cups are
used, and if these rotors or safety cups are opened only in a biological
safety cabinet.
2.
Face protection (goggles, mask, face shield or other splatter guards) is required to
prevent splashes or sprays of infectious or other hazardous materials to the face,
when the microorganisms must be manipulated outside the BSC.
3.
Protective laboratory coats, gowns, smocks, or uniforms designated for lab use are
worn while in the laboratory. This protective clothing is removed and left in the
laboratory before leaving for non-laboratory areas (e.g., cafeteria, library, and
administrative offices). All protective clothing is either disposed of in the
laboratory or laundered by the institution; personnel should never take it home.
4.
Gloves are required when handling infected animals and when hands may contact
infectious materials, contaminated surfaces or equipment. Wearing two pairs of
gloves may be appropriate; if a spill or splatter occurs, the hand will be protected
after the contaminated glove is removed. Gloves are disposed of when
Biosafety Manual
19
contaminated, removed when work with infectious materials is completed, and are
never worn outside the laboratory. Disposable gloves are not washed or reused.
D.
Laboratory Facilities (Secondary Barriers)
1.
Each laboratory contains a sink for hand washing.
2.
The laboratory is designed so that it can be easily cleaned. Walls and floors should
be smooth, hard and constructed of water impervious material that will stand up to
harsh disinfectants. Carpeting is not allowed in laboratory facilities.
3.
Bench tops should be smooth, hard and impervious to water and resistant to acids,
alkalis, organic solvents, and moderate heat.
4.
Laboratory furniture is sturdy, and spaces between benches, cabinets, and
equipment are accessible for cleaning. No cloth or fabric seating is permitted.
5.
If the laboratory has windows that open, they are fitted with fly screens.
6.
A method for decontamination of infectious or regulated laboratory wastes is
available (e.g., autoclave, chemical disinfection, incinerator, or other approved
decontamination system).
7.
An eyewash/safety shower facility is readily available.
8.
All laboratories require single pass air that is not then recirculated to any other
area of the facility. Laboratories should be negative pressure to surrounding areas
to prevent accidental spread of potentially infectious or recombinant agents.
9.
Install biological safety cabinets in such a manner that fluctuations of the room
supply and exhaust air do not cause the biological safety cabinets to operate
outside their parameters for containment. Locate biological safety cabinets away
from doors, from windows that can be opened, from heavily traveled laboratory
areas, and from other potentially disruptive equipment so as to maintain the
biological safety cabinets’ air flow parameters for containment.
Biosafety Level 3
Biosafety Level 3 is applicable to clinical, diagnostic, teaching, research, or production facilities
in which work is done with indigenous or exotic agents which may cause serious or potentially
lethal disease as a result of exposure by the inhalation route. 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 devices, or by personnel wearing
appropriate personal protective clothing and equipment. The laboratory has special engineering
and design features.
It is recognized, however, that many existing facilities may not have all the facility safeguards
recommended for Biosafety Level 3 (e.g., access zone, sealed penetrations, directional airflow,
etc.). In these circumstances, acceptable safety may be achieved for routine or repetitive
operations (e.g. diagnostic procedures involving the propagation of an agent for identification,
typing, and susceptibility testing) in Biosafety Level 2 facilities. However, the recommended
20
Biosafety Manual
Standard Microbiological Practices, Special Practices, and Safety Equipment for Biosafety Level
3 must be rigorously followed. The decision to implement this modification of Biosafety Level 3
recommendations should be made only by the lab director.
The following standard and special safety practices, equipment, and facilities apply to the use of
agents assigned to Biosafety Level 3:
A.
Standard Microbiological Practices
1.
Access to the laboratory is limited or restricted at the discretion of the laboratory
director when experiments are in progress.
2.
Persons wash their hands after handling infectious materials and animals, after
removing gloves, and when they leave the laboratory.
3.
Eating, drinking, the use of tobacco products, handling contact lenses and
applying cosmetics or lip balm are not permitted in the laboratory. Persons who
wear contact lenses in laboratories should also wear goggles or a face shield.
Food is stored outside the work area in cabinets or refrigerators designated for this
purpose only.
4.
Mouth pipetting is prohibited; mechanical pipetting devices are used.
5.
Policies for the safe handling of sharps are instituted.
6.
All procedures are performed carefully to minimize the creation of aerosols.
7.
Work surfaces are decontaminated at least once a day and immediately after any
spill of viable material.
8.
All cultures, stocks, and regulated wastes are decontaminated before disposal by
an approved decontamination method, such as autoclaving. Materials to be
decontaminated outside of the immediate laboratory are to be placed in a durable,
leak-proof container and closed for transport from the laboratory. Infectious
wastes from BSL-3 laboratories must be decontaminated before removal for offsite disposal.
9.
An insect and rodent control program is in effect.
10.
Policies for the safe handling of sharps are instituted.
B.
Special Practices
1.
Laboratory doors are kept closed at all times.
2.
The laboratory director controls access to the laboratory and restricts access to
persons whose presence is required for program or support purposes. For
example, persons who are immunocompromised or immunosuppressed, or for
whom infection may be unusually hazardous, are not allowed in the laboratory or
animal rooms. The director has the final responsibility for assessing each
circumstance and determining who may enter or work in the laboratory. No
minors are allowed in level 3 laboratories.
3.
The laboratory director establishes policies and procedures whereby only persons
who have been advised of the potential biohazard, who meet any specific entry
requirements (e.g., immunizations), and who comply with all entry and exit
procedures, enter the laboratory or animal rooms.
Biosafety Manual
21
4.
5.
6.
7.
8.
9.
10.
22
When infectious materials or infected animals are present in the laboratory or
containment module, a hazard warning sign incorporating the universal biohazard
symbol, is posted on all laboratory and animal room access doors. The hazard
warning sign identifies the agent, lists the name and telephone number of the
laboratory director or other responsible person(s), and indicates any special
requirements for entering the laboratory, such as the need for immunizations,
respirators, or other personal protective measures.
Laboratory personnel receive the appropriate immunizations or tests for the agents
handled or potentially present in the laboratory (e.g., hepatitis B vaccine or TB
skin testing and periodic testing).
Baseline serum samples are collected and stored for all laboratory and other atrisk personnel. Additional serum specimens may be collected periodically,
depending on the agents handled or the function of the laboratory.
A Biosafety Manual specific to the laboratory is prepared or adopted. Personnel
are advised of special hazards and are required to read and to follow instructions
on practices and procedures. An SOP manual is prepared to cover all procedures
and activities.
Laboratory personnel receive appropriate training on the potential hazards
associated with the work involved, the necessary precautions to prevent
exposures, and the exposure evaluation procedures. Personnel receive annual
updates, or additional training as necessary for procedural changes.
The laboratory director is responsible for insuring that all personnel demonstrate
proficiency in standard microbiological practices and techniques, and in the
practice and operations specific to the laboratory facility, before working with
organisms at Biosafety Level 3. This might include prior experience in handling
human pathogens or cell cultures, or a specific training program provided by the
laboratory director or other competent scientist proficient in safe microbiological
practices and techniques.
A high degree of precaution must always be taken with all contaminated sharp
items, including needles and syringes, slides, pipettes, capillary tubes, and
scalpels. Needles and syringes or other sharp instruments should be restricted in
the laboratory for use only when there is no alternative, such as parenteral
injection, phlebotomy, or aspiration of fluids from laboratory animal and
diaphragm bottles. Plastic ware should be substituted for glassware whenever
possible.
a.
Only needle-locking syringes or disposable syringe needle units (i.e.,
needle is integral to the syringe) are used for injection or aspiration of
infectious materials. Used disposable needles must not be bent, sheared,
broken, recapped, removed from disposable syringes, or otherwise
manipulated by hand before disposal; rather, they must be carefully placed
in conveniently located puncture-resistant containers used for sharps
disposal. Non-disposable sharps must be placed in a hard-walled
container for transport to a processing area for decontamination, preferably
by autoclaving.
Biosafety Manual
b.
11.
12.
13.
14.
15.
16.
17.
C.
Syringes that re-sheathe the needle, needle-less systems, and other safe
devices should be used when appropriate.
c.
Broken glassware must not be handled directly by hand, but must be
removed by mechanical means such as a brush and dustpan, tongs, or
forceps. Containers of contaminated needles, sharp equipment, and
broken glass should be decontaminated before disposal, in accordance
with any local, state, or federal regulations.
All manipulations involving infectious materials are conducted in biological
safety cabinets or other physical containment devices within the containment
module. No work in open vessels is conducted on the open bench.
Laboratory equipment and work surfaces should be decontaminated with an
appropriate disinfectant on a routine basis, after work with infectious materials is
finished. It must be cleaned and decontaminated after overt spills, splashes, or
other contamination with infectious materials. Contaminated equipment should
also be decontaminated before it is sent for repair or maintenance. An Equipment
Release Form, available on the EH&S website, should be used to assure that
equipment has been properly decontaminated prior to servicing. In addition, it
must be packaged for transport in accordance with applicable local, state, or
federal regulations, before removal from the facility. Plastic-backed paper
toweling can be used on non-perforated work surfaces within biological safety
cabinets to facilitate clean up.
Cultures, tissues, or specimens of body fluids are placed in a container that
prevents leakage during collection, handling, processing, storage, transport, or
shipping.
All potentially contaminated materials (e.g., gloves, lab coats, etc.) from
laboratories or animal rooms are decontaminated before disposal or reuse.
Spills of infectious materials are decontaminated, contained and cleaned up by
appropriate professional staff, or others properly trained and equipped to work
with concentrated infectious material.
Spills and accidents that result in overt or potential exposures to infectious
materials are immediately reported to the laboratory director. Appropriate
medical evaluation, surveillance, and treatment are provided and written records
are maintained.
Animals and plants not related to the work being conducted are not permitted in
the laboratory.
Safety Equipment (Primary Barriers)
1.
Properly maintained biological safety cabinets are used (Class II or III) for all
manipulation of infectious materials. Biological Safety Cabinets (BSC) should be
located away from doors, air supplies and other heavily traveled areas. Biosafety
Cabinets must be certified at least yearly.
2.
Outside of a BSC, appropriate combinations of personal protective equipment are
used (e.g., special protective clothing, masks, gloves, face protection, or
respirators) in combination with physical containment devices (e.g., centrifuge
safety cups, sealed centrifuge rotors, or containment caging for animals).
Biosafety Manual
23
3.
4.
5.
6.
7.
D.
24
Biological Safety Cabinets must be used for manipulations of cultures and clinical
or environmental materials that may be a source of infectious aerosols. The
aerosol challenge of experimental animals; harvesting of tissues or fluids from
infected animals, and embryonated eggs, and necropsy of infected animals also
require the use of BSCs.
Face protection (goggles and mask, or face shield) is worn for manipulations of
infectious materials outside of a biological safety cabinet.
Respiratory protection is worn when aerosols cannot be safety contained (e.g.,
outside of a biological safety cabinet), and in rooms containing infected animals.
Protective laboratory clothing such as solid-front or wrap-around gowns, scrub
suits, or coveralls must be worn in, and not worn outside, the laboratory.
Reusable laboratory clothing is to be decontaminated before being laundered.
Change protective clothing immediately if contaminated.
Gloves must be worn when handling infected animals and when hands may
contact infectious materials and contaminated surfaces or equipment. Disposable
gloves should be discarded when contaminated, and never washed for reuse.
Always wash hands between glove changes.
Laboratory Facilities (Secondary Barriers)
1.
The laboratory is separated from areas that are open to unrestricted traffic flow
within the building. Passage through two sets of self-closing doors is the basic
requirement for entry into the laboratory from access corridors or other contiguous
areas. A clothes change room (shower optional) may be included in the
passageway.
2.
Each laboratory contains a sink for hand washing. The sink is foot, elbow, or
automatically operated and is located near the laboratory exit door.
3.
The interior surfaces of walls, floors, and ceilings are smooth, hard, impervious,
and water-resistant so that they can be easily cleaned and capable of disinfection.
Penetrations around these surfaces are sealed or capable of being sealed to
facilitate decontamination. Floors should be monolithic and coved to the walls.
4.
Bench tops are smooth, hard and impervious to water and resistant to acids,
alkalis, organic solvents, and moderate heat.
5.
Laboratory furniture is sturdy, and spaces between benches, cabinets, and
equipment are accessible for cleaning. No fabric materials are allowed.
6.
Windows in the laboratory are closed and sealed.
7.
A method for decontaminating all laboratory wastes is available, preferably within
the laboratory (i.e., autoclave, chemical disinfection, incineration, or other
approved decontamination method).
8.
A ducted exhaust air ventilation system is provided. This system creates
directional airflow that draws air from “clean” areas into the laboratory toward
“contaminated” areas. The exhaust air is not recirculated to any other area of the
building, and, depending on risk assessment, discharged to the outside through a
HEPA filtration system. The outside exhaust must be dispersed away from
occupied areas and air intakes. Laboratory personnel must verify that the
Biosafety Manual
9.
10.
11.
12.
13.
14.
15.
16.
direction of the airflow (into the laboratory) is proper. Visual monitoring devices,
such as digital readout or maghihelic gauges are recommended.
The High Efficiency Particulate Air (HEPA)-filtered exhaust air from Class II or
Class III biological safety cabinets is discharged directly to the outside or through
the building exhaust system. If the HEPA-filtered exhaust air from Class II Type
A biological safety cabinets is to be discharged to the outside through the building
exhaust air system, it is connected to this system in a manner (e.g., thimble unit
connection) that avoids any interference with the air balance of the cabinets or
building exhaust system. Exhaust air from Class II type A biological safety
cabinets may be exhausted into the laboratory if the cabinet is tested and certified
at least every twelve months. Class III biosafety cabinets must be hard ducted to
the building exhaust system.
Continuous flow centrifuges or other equipment that may produce aerosols are
contained in devices that exhaust air through HEPA filters before discharge into
the laboratory.
Vacuum lines are protected with liquid disinfectant traps or HEPA filters, or their
equivalent, which are routinely maintained and replaced as needed.
An eyewash/safety shower is readily available within the BSL-3 suite.
Biosafety Level 3 facility design and operational procedures must be documented.
The facility must be tested for verification that the design and operational
parameters have been met prior to operation. Facilities should be reverified/certified at least annually (HEPAs and air systems).
Additional environmental protection (e.g. personnel showers, containment of
other piped services and the provision for effluent decontamination) should be
added when recommended by the agent summary statement, as determined by risk
assessment, the site conditions or other applicable federal, state, or local
regulations.
Level 3 laboratories and animal areas are audited by The EH&S Biosafety Office
quarterly.
Openings such as around ducts, spaces between doors and frames and other
penetrations must be capable of being sealed to facilitate decontamination.
Animal Biosafety Level 1 (ABSL-1)
Animal Biosafety Level 1 (ABSL-1) is
suitable for work involving well
characterized agents that are not known to
cause disease in healthy adult humans, and
that are of minimal potential hazard to
laboratory personnel and the environment.
Biosafety Manual
25
A.
Standard Practices
1.
The animal facility director establishes polices, procedures, and protocols for
emergency situations. Each project is subject to pre-approval by the Institutional
Animal Care and Use Committee (IACUC) and the Institutional Biosafety
Committee (IBC). Any special practices are approved at this time.
2.
Only those persons required for program or support purposes are authorized to
enter the facility. Before entering, persons are advised of the potential biohazards
and are instructed on the appropriate safeguards.
3.
An appropriate medical surveillance program is in place.
4.
A safety manual is prepared or adopted. Personnel are advised of special hazards,
and are required to read and follow instructions on practices and procedures.
5.
Eating, drinking, smoking, handling contact lenses, applying cosmetics, and
storing food for human use should only be done in designated areas and are not
permitted in animal or procedure rooms.
6.
All procedures are carefully performed to minimize the creation of aerosols or
splatters.
7.
Work surfaces are decontaminated after use or after any spill of viable materials.
8.
All wastes from the animal room (including animal tissues, carcasses, and
contaminated bedding) are transported from the animal room in leak-proof,
covered containers for appropriate disposal in compliance with applicable
institutional or local requirements. As a means of incineration is not available on
the NAU Campus, all animal wastes must be packaged and shrouded and
delivered to the Vivarium Manager at the Biological Sciences Annex.
9.
Policies for the safe handling of sharps are instituted.
10.
Personnel wash their hands after handling cultures and animals, after removing
gloves, and before leaving the animal facility.
11.
A biohazard sign must be posted on the entrance to the animal room whenever
infectious agents are present. The hazard warning sign identifies the infectious
agent(s) in use, lists the name and telephone number of the responsible person(s),
and indicates the special requirements for entering the animal room (e.g., the need
for immunizations and respirators).
12.
An insect and rodent control program is in effect.
B.
Special Practices: None
C.
Safety Equipment (Primary Barriers):
1.
The wearing of laboratory coats, gowns, and/or uniforms in the facility is
recommended. Laboratory coats remain in the animal room. Gowns and
uniforms are not worn outside the facility.
2.
Persons having contact with non-human primates should assess their risk of
mucous membrane exposure and wear appropriate eye and face protection.
D.
Facilities (Secondary Barriers)
1.
The animal facility is separated from areas that are open to unrestricted personnel
traffic within the building.
2.
External facility doors are self-closing and self-locking. Doors to animal rooms
open inward, are self-closing, and are kept closed when experimental animals are
26
Biosafety Manual
3.
4.
5.
6.
7.
8.
9.
10.
present. Cubicle room inner doors may open outward or be horizontal or vertical
sliding.
The animal facility is designed, constructed, and maintained to facilitate cleaning
and housekeeping. The interior surfaces (walls, floors, and ceilings) are water
resistant.
Internal facility appurtenances, such as light fixtures, air ducts, and utility pipes,
are arranged to minimize horizontal surface areas.
Windows are not recommended. Any windows must be resistant to breakage.
Where possible, windows should be sealed. If the animal facility has windows that
open, they are fitted with fly screens.
If floor drains are provided, the traps are always filled with water and/or an
appropriate disinfectant.
Ventilation should be provided in accordance with the Guide for Care and Use of
Laboratory Animals, latest edition. No recirculation of exhaust air should occur. It
is recommended that animal rooms maintain negative pressure compared to
adjoining hallways.
The facility has a hand washing sink.
Cages are washed manually or in a cage washer. The mechanical cage washer
should have a final rinse temperature of at least 180F.
Illumination is adequate for all activities, avoiding reflections and glare that could
impede vision.
Animal Biosafety Level 2 (ABSL-2)
Animal Biosafety Level 2 involves practices for work with those
agents associated with human disease. It addresses hazards from
ingestion as well as from percutaneous and mucous membrane
exposure. ABSL-2 builds upon the practices, procedures,
containment equipment, and facility requirements of ABSL-1.
A.
Standard Practices
1.
Aside from the standard policies, procedures, and protocols for emergency
situations established by the facility director, appropriate special policies and
procedures should be developed as needed and approved by the Institutional
Animal Care and Use Committee (IACUC) and the Institutional Biosafety
Committee (IBC).
2.
Access to the animal room is limited to the fewest number of individuals possible.
Personnel who must enter the room for program or service purposes when work is
in progress are advised of the potential hazard.
Biosafety Manual
27
3.
4.
5.
6.
7.
8.
9.
10.
11.
12.
B.
28
An appropriate medical surveillance program is in place. All personnel receive
appropriate immunizations or tests for the agents handled or potentially present
(e.g., hepatitis B vaccine, TB skin testing). When appropriate, a serum
surveillance system should be implemented.
A Biosafety Manual is prepared or adopted. Personnel are advised of special
hazards, and are required to read and follow instructions on practices and
procedures.
Eating, drinking, smoking, handling contact lenses, applying cosmetics, and
storing food for human use should only be done in designated areas and are not
permitted in animal or procedure rooms.
All procedures are carefully performed to minimize the creation of aerosols or
splatters.
Equipment and work surfaces in the room are routinely decontaminated with an
effective disinfectant after work with the infectious agent, and especially after
overt spills, splashes, or other contamination by infectious materials.
All infectious samples are collected, labeled, transported, and processed in a
manner that contains and prevents transmission of the agent(s). All wastes from
the animal room (including animal tissues, carcasses, contaminated bedding,
unused feed, sharps, and other refuse) are transported from the animal room in
leak-proof, covered containers for appropriate disposal in compliance with
applicable institutional or local requirements. The outer surface of the containers
is disinfected prior to moving the material. As a means of incineration is not
available on the NAU Campus, all animal wastes must be packaged and shrouded
and delivered to the Vivarium Manager at the Biological Sciences Annex.
Policies for the safe handling of sharps are instituted:
a.
Needles and syringes or other sharp instruments are restricted for
use in the animal facility only when there is no alternative, such as
for parenteral injection, blood collection, or aspiration of fluids
from laboratory animals and diaphragm bottles.
b.
Syringes that re-sheathe the needle, needle-less systems, and other
safe devices should be used when appropriate.
c.
Plastic-ware should be substituted for glassware whenever
possible.
Personnel wash their hands after handling cultures and animals, after removing
gloves, and before leaving the animal facility.
A biohazard sign must be posted on the entrance to the animal room whenever
infectious agents are present. The hazard warning sign identifies the infectious
agent(s) in use, lists the name and telephone number of the responsible person(s),
and indicates the special requirements (e.g., the need for immunizations and
respirators) for entering the animal room.
An insect and rodent control program is in effect.
Special Practices
1.
Animal care laboratory and support personnel receive appropriate training on the
potential hazards associated with the work involved, the necessary precautions to
Biosafety Manual
2.
3.
4.
prevent exposures, and the exposure evaluation procedures. Personnel receive
annual updates, or additional training as necessary for procedural or policy
changes. Records of all training provided are maintained. In general, persons who
may be at increased risk of acquiring infection, or for whom infection might be
unusually hazardous, are not allowed in the animal facility unless special
procedures can eliminate the extra risk.
Only animals used for the experiment(s) are allowed in the room.
All equipment must be appropriately decontaminated prior to removal from the
room.
Spills and accidents which result in overt exposures to infectious materials must
be immediately reported to the facility director. Medical evaluation, surveillance,
and treatment are provided as appropriate and written records are maintained.
C.
Safety Equipment (Primary Barriers)
1.
Gowns, uniforms, or laboratory coats are worn while in the animal room. The
laboratory coat is removed and left in the animal room. Gowns, uniforms, and
laboratory coats are removed before leaving the animal facility. Gloves are worn
when handling infected animals and when skin contact with infectious materials is
unavoidable.
2.
Personal protective equipment is used based on risk assessment determinations.
Appropriate face/eye and respiratory protection is worn by all personnel entering
animal rooms that house nonhuman primates.
3.
Biological safety cabinets, other physical containment devices, and/or personal
protective equipment (e.g., respirators, face shields) are used whenever
conducting procedures with a high potential for creating aerosols. These include
necropsy of infected animals, harvesting of tissues or fluids from infected animals
or eggs, or intranasal inoculation of animals.
4.
When needed, animals are housed in primary biosafety containment equipment
appropriate for the animal species. Filter top cages are always handled in properly
designed and operating animal bio-containment cabinets recommended for
rodents.
D.
Facilities (Secondary Barriers)
1.
The animal facility is separated from areas that are open to unrestricted personnel
traffic within the building.
2.
Access to the facility is limited by secure locked doors. External doors are selfclosing and self-locking. Doors to animal rooms open inward, are self-closing,
and are kept closed when experimental animals are present. Cubicle room inner
doors may open outward or be horizontal or vertical sliding.
3.
The animal facility is designed, constructed, and maintained to facilitate cleaning
and housekeeping. The interior surfaces (walls, floors, and ceilings) are smooth,
hard and impervious.
4.
Internal facility appurtenances, such as light fixtures, air ducts, and utility pipes,
are arranged to minimize horizontal surface areas.
Biosafety Manual
29
5.
6.
7.
8.
9.
10.
11.
Any windows must be resistant to breakage. Where possible, windows should be
sealed. If the animal facility has windows that open, they are fitted with fly
screens.
If floor drains are provided, the traps are always filled with an appropriate
disinfectant.
Exhaust air is discharged to the outside without being recirculated to other rooms.
Ventilation should be provided in accordance with criteria from Guide for Care
and Use of Laboratory Animals, latest edition. The direction of airflow in the
animal facility is inward; animal rooms should maintain negative pressure
compared to adjoining hallways.
Cages are washed manually or in an appropriate cage washer. The mechanical
cage washer should have a final rinse temperature of at least 180F.
An autoclave is available in the animal facility to decontaminate infectious waste.
A hand washing sink is in the animal room where infected animals are housed, as
well as elsewhere in the facility.
Illumination is adequate for all activities, avoiding reflections and glare that could
impede vision.
Animal Biosafety Level 3 (ABSL-3)
Animal Biosafety Level 3 involves practices suitable for work with animals infected with
indigenous or exotic agents that present the potential of aerosol transmission and of causing
serious or potentially lethal disease. ABSL-3 builds upon the standard practices, procedures,
containment equipment, and facility requirements of ABSL-2.
A.
30
Standard Practices
1.
Aside from the standard policies, procedures, and protocols for emergency
situations established by the facility director, appropriate special policies and
procedures should be developed as needed and approved by the Institutional
Animal Care and Use Committee (IACUC) and the Institutional Biosafety
Committee (IBC).
2.
The laboratory or animal facility director limits access to the animal room to the
fewest number of individuals possible. Personnel who must enter the room for
program or service purposes when work is in progress are advised of the potential
hazard.
3.
An appropriate medical surveillance program is in place. All personnel receive
appropriate immunizations or tests for the agents handled or potentially present
(e.g., hepatitis B vaccine, TB skin testing). When appropriate, a serum
surveillance system should be implemented. In general, persons who may be at
increased risk of acquiring infection, or for whom infection might have serious
consequences, are not allowed in the animal facility unless special procedures can
eliminate the extra risk. Assessment should be made by the occupational health
physician.
Biosafety Manual
4.
5.
6.
7.
8.
9.
10.
11.
12.
13.
14.
B.
A Biosafety Manual is prepared or adopted. Personnel are advised of special
hazards, and are required to read and follow instructions on practices and
procedures.
Eating, drinking, smoking, handling contact lenses, applying cosmetics, and
storing food for human use should be done only in designated areas and are not
permitted in animal or procedure rooms.
All procedures are carefully performed to minimize the creation of aerosols or
splatters.
Equipment and work surfaces in the room are routinely decontaminated with an
effective disinfectant after work with the infectious agent, and especially after
overt spills, splashes, or other contamination by infectious materials.
All wastes from the animal room (including animal tissues, carcasses,
contaminated bedding, unused feed, sharps, and other refuse animal tissues) are
transported from the animal room in leak-proof, covered containers for
appropriate disposal in compliance with applicable institutional or local
requirements. The outer surface of the containers is disinfected prior to moving
the material. As a means of incineration is not available on the NAU Campus, all
animal wastes must be packaged and shrouded and delivered to the Vivarium
Manager at the Biological Sciences Annex. (see Special Practices #3 below).
Policies for the safe handling of sharps are instituted:
a.
Needles and syringes or other sharp instruments are restricted in the
animal facility for use only when there is no alternative, such as for
parenteral injection, blood collection, or aspiration of fluids from
laboratory animals and diaphragm bottles.
b.
Syringes that re-sheathe the needle, needle-less systems, and other safety
devices should be used when appropriate.
c.
Plastic ware should be substituted for glassware whenever possible.
Personnel wash their hands after handling cultures and animals, after removing
gloves, and before leaving the animal facility.
A biohazard sign must be posted on the entrance to the animal room whenever
infectious agents are present. The hazard warning sign identifies the infectious
agent(s) in use, lists the name and telephone number of the responsible person(s),
and indicates the special requirements for entering the animal room (e.g., the need
for immunizations and respirators).
All infectious samples are collected, labeled, transported, and processed in a
manner that contains and prevents transmission of the agent(s).
Laboratory and support personnel receive appropriate training on the potential
hazards associated with the work involved, the necessary precautions to prevent
exposures, and the exposure evaluation procedures. As necessary, personnel
receive updates and/or additional training on procedural or policy changes.
Records of all training provided are maintained.
An insect and rodent control program is in effect.
Special Practices
Biosafety Manual
31
1.
2.
3.
C.
Safety Equipment (Primary Barriers)
1.
Uniforms or scrub suits are worn by personnel entering the
animal room. Wrap-around or solid-front gowns should be
worn over this clothing. Front-button laboratory coats are
unsuitable. The gown must be removed and left in the
animal room. Before leaving the animal facility, scrub
suits and uniforms are removed and appropriately
contained and decontaminated prior to laundering or
disposal.
2.
Personal protective equipment used is based on risk
assessment determinations.
a.
Personal protective equipment is used for all
activities involving manipulations of infectious
material or infected animals.
b.
Personnel wear gloves when handling infected
animals. Gloves are removed aseptically and
autoclaved with other animal room wastes before
disposal.
c.
Appropriate face/eye and respiratory protection (e.g., respirators and face
shields) is worn by all personnel entering animal rooms.
d.
Boots, shoe covers, or other protective footwear, and disinfectant foot
baths are available and used where indicated.
3.
4.
32
Cages are autoclaved or thoroughly decontaminated before bedding is removed
and before they are cleaned and washed. Equipment must be decontaminated
according to any local, state, or federal regulations before being packaged for
transport or removal from the facility for repair or maintenance.
A spill procedure is developed and posted. Only personnel properly trained and
equipped to work with infectious materials are to clean up spills. Spills and
accidents that result in overt exposures to infectious materials must be
immediately reported to the facility director. Medical evaluation, surveillance, and
treatment are provided as appropriate and written records are maintained.
Materials not related to the experiment (e.g., plants, animals) are not permitted in
the animal room.
The risk of infectious aerosols from infected animals or their bedding also can be
reduced if animals are housed in containment caging systems, such as open cages
placed in inward flow ventilated enclosures (e.g., laminar flow cabinets), solid
wall and bottom cages covered with filter bonnets, or other equivalent primary
containment systems.
Biological safety cabinets and other physical containment devices are used
whenever conducting procedures with a potential for creating aerosols. These
include necropsy of infected animals, harvesting of tissues or fluids from infected
animals or eggs, or intranasal inoculation of animals. At ABSL-3, all work should
Biosafety Manual
be done in a primary barrier; otherwise respirators should be worn by personnel in
the room.
D.
Facilities (Secondary Barriers)
1.
The animal facility is separated from areas that are open to unrestricted personnel
traffic within the building.
2.
Access to the facility is limited by a self-closing and self-locking door. This
exterior entry door may be controlled by a key lock, card key, biometrics, or
proximity reader. Entry into the animal room is via a double-door entry which
includes a change room and shower(s). An additional double-door access (airlock) or double-doored autoclave may be provided for movement of supplies and
wastes into and out of the facility, respectively. Doors to animal rooms open
inward and are self-closing. Doors to cubicles inside an animal room may open
outward or slide horizontally or vertically.
3.
The animal facility is designed, constructed, and maintained to facilitate cleaning
and housekeeping. The interior surfaces (walls, floors, and ceilings) are smooth,
hard, impervious and capable of harsh disinfection. Penetrations in floors, walls
and ceiling surfaces are sealed and openings around ducts and the spaces between
doors and frames are capable of being sealed to facilitate decontamination.
4.
A hands-free or automatically operated hand washing sink is provided in each
animal room near the exit door. The sink trap is filled with an appropriate
disinfectant after each use.
5.
Internal facility appurtenances, such as light fixtures, air ducts, and utility pipes,
are arranged to minimize horizontal surface areas.
6.
Windows are not recommended. Any windows must be resistant to breakage and
must be sealed. Direct viewing of animals is prohibited.
7.
If floor drains are provided, they are always filled with an appropriate disinfectant.
8.
Ventilation should be provided in accordance with criteria from the Guide for
Care and Use of Laboratory Animals, latest edition. A ducted exhaust air
ventilation system is provided. This system creates directional airflow which
draws air into the laboratory from "clean" areas and toward "contaminated" areas.
The exhaust air is not recirculated to any other area of the building. Filtration and
other treatments of the exhaust air may not be required, but should be considered
based on site requirements, and specific agent manipulations and use conditions.
The exhaust must be dispersed away from occupied areas and air intakes, or the
exhaust must be HEPA-filtered. Personnel must verify that the direction of the
airflow (into the animal areas) is proper. It is recommended that a visual
monitoring device that indicates and confirms directional inward airflow be
provided at the animal room entry. Consideration should be given to installing an
HVAC control system to prevent sustained positive pressurization of the animal
spaces. Audible alarms should be considered to notify personnel of HVAC system
failure.
9.
HEPA-filtered exhaust air from a Class II type A biological safety cabinet can be
exhausted into the animal room if the cabinet is tested and certified at least
annually. When exhaust air from Class II type A safety cabinets is to be
Biosafety Manual
33
10.
11.
12.
13.
14.
15.
34
discharged to the outside through the building exhaust air system, the cabinets
must be connected in a manner that avoids any interference with the air balance of
the cabinets or the building exhaust system (e.g., a thimble unit, or an air gap
between the cabinet exhaust and the exhaust duct). When Class III biological
safety cabinets are used, they should be directly connected (hard ducted) to the
exhaust system
Cages are washed in a cage washer. The mechanical cage washer has a final rinse
temperature of at least 180F.
An autoclave is available which is convenient to (preferably within) the animal
rooms where the biohazard is contained. The autoclave is utilized to
decontaminate infectious waste before moving it to other areas of the facility. A
Pass Through autoclave is recommended.
If vacuum service (i.e., central or local) is provided, each service connection
should be fitted with liquid disinfectant traps and an in-line HEPA filter, placed as
near as practicable to each use point or service cock. Filters are installed to permit
in-place decontamination and replacement.
Illumination is adequate for all activities, avoiding reflections and glare that could
impede vision.
The completed Biosafety Level 3 facility design and operational procedures must
be documented. The facility must be tested for verification that the design and
operational parameters have been met prior to operation. Facilities should be reverified at least annually against these procedures as modified by operational
experience.
Additional environmental protection (e.g., personnel showers, HEPA filtration of
exhaust air, containment of other piped services, and the provision of effluent
decontamination) should be considered if recommended by the agent summary
statement, as determined by risk assessment of the site conditions, or other
applicable federal, state, or local regulations. If USDA oversight or funding is
involved, HEPA filtered exhaust, personnel shower in/shower out and a Pass
Through autoclave will be required.
Biosafety Manual
Table 1: Summary of Recommended Biosafety Levels for Infectious Agents
BSL
Agents
1
Not known to consistently
cause disease in healthy
adults
Standard Microbiological Practices
None required
Open bench top
Sink for hand washing
2
Associated with human
disease, hazard =
percutaneous injury,
ingestion, mucous
membrane exposure
BSL-1 practice plus:
 Limited access
 Biohazard warning signs
 “Sharps” precautions
 Biosafety manual defining any
needed waste decontamination
or medical surveillance
policies
Primary barriers = Class I or II
BSCs or other physical containment
devices used for all manipulations
of agents that cause splashes or
aerosols of infectious materials;
PPE; laboratory coats; gloves; face
protection as needed
BSL-1 plus:
 Autoclave available
 Single pass air with no
recirculation
 Hard, smooth,
impervious floor, walls,
and countertops
3
Indigenous or exotic
agents with potential for
aerosol transmission;
disease may have serious
or lethal consequences
BSL-2 practice plus:
 Controlled access
 Decontamination of all waste
 Decontamination of lab
clothing before laundering
 Baseline serum
 Decontamination of all
effluent
Primary barriers = Class I or II
BSCs or other physical containment
devices for all open manipulations
of agents; PPE; protective lab
clothing; gloves; respiratory
protection as needed
BSL-2 plus:
 Physical separation from
access corridors
 Self-closing, doubledoor access
 Exhausted air not recirculated
 Negative airflow into
laboratory (single pass)
 No floor drains
 Hard, smooth,
impervious ceilings
4
Dangerous/exotic agents
which pose high risk of
life-threatening disease,
aerosol-transmitted lab
infections; or related
agents with unknown risk
of transmission
BSL-3 practice plus:
 Clothing change before
entering
 Shower on exit
 All material decontaminated
on exit from facility
Primary barriers = All procedures
conducted in Class III BSCs or
Class I or II BSCs in combination
with full-body air-supplied, positive
pressure personnel suit
BSL-3 plus:
 Separate building or
isolated zone
 Dedicated supply and
exhaust, vacuum, and
decontamination systems
 Other requirements
outlined in the text
Biosafety Manual
Practices
Safety Equipment
(Primary Barriers)
35
Facilities
(Secondary Barriers)
Table 2: Summary of Recommended Biosafety Levels for Activities in Which Experimentally or Naturally Infected
Vertebrate Animals Are Used
ABSL
Agents
Practices
Safety Equipment
(Primary Barriers)
Facilities
(Secondary Barriers)
1
Not known to
consistently cause
disease in healthy
adults
Standard animal care and management
practices, including appropriate medical
surveillance programs
As required for normal care of each
species
Standard animal facility
 No recirculation of exhaust air
 Directional air flow recommended
 Hand washing sink recommended
2
Associated with
human disease,
hazard =
percutaneous injury,
ingestion, mucous
membrane exposure
ABSL-1 practice plus:
 Limited access
 Biohazard warning signs
 “Sharps” precautions
 Decontamination of all
infectious wastes and of animal
cages prior to washing
ABSL-1 equipment plus barriers:
containment equipment appropriate for
animal species; PPE; laboratory coats,
gloves, face and respiratory protection
as needed.
ABSL-1 facility plus:
 Autoclave available
 Hand washing sink available in
animal room
 Mechanical cage washer used
 Hard, smooth, impervious walls,
floors, ceilings and countertops
3
Indigenous or exotic
agents with potential
for aerosol
transmission; disease
may have serious or
lethal consequences
ABSL-2 practice plus:
 Controlled access
 Decontamination of all waste
 Cages decontaminated before
bedding removed
 Disinfectant foot bath as
needed
 Decontamination of all effluent
ABSL-2 equipment plus:
 Containment equipment for
housing animals and cage
dumping activities
 Class I or II BSCs available
for manipulative
procedures(inoculation,
necropsy) that may create
infectious aerosols. PPE:
appropriate respiratory
protection
ABSL-2 facility plus:
 Physical separation from access
corridors
 Self-closing, double-door access
 Sealed penetrations
 Sealed windows
 Autoclave available in facility
4
Dangerous/exotic
agents which pose
high risk of lifethreatening disease,
aerosol-transmitted
lab infections; or
related agents with
unknown risk of
transmission
ABSL-3 practice plus:
 Entrance through change room
where personal clothing is
removed and laboratory
clothing is put on; shower on
exit
 All wastes are decontaminated
before removal from facility
ABSL-3 equipment plus maximum
containment equipment (i.e. Class III
BSC or partial containment equipment
in combination with full body, airsupplied positive-pressure personnel
suit) used for all procedures and
activities
ABSL-3 facility plus:
 Separate building or isolated zone
 Dedicated air supply and exhaust,
vacuum and decontamination
systems
 Other requirements outlined in the
text
36
Biosafety Manual
Agents List
The following agents have been listed according to the most appropriate Biological Safety Level to
be used. The lists presented on the following pages are based upon the most recent information
available in the “Risk Group Classifications for Infectious Agents” section of the American
Biological Safety Association (ABSA) Website. Every attempt to provide accurate classification for
the agents listed has been made. As this is not a complete list of infectious agents, and these
classifications are subject to change, please refer to the following website: http://www.phacaspc.gc.ca/lab-bio/res/psds-ftss/index-eng.php, for classification of other agents, and the most up to
date information for those listed.
Please note that Biological Safety Levels are not inherent to an agent but are performance
recommendations and should be chosen after a risk assessment is completed. A proper risk
assessment takes into account the characteristics of the agent involved, the activities to be
performed, and the environment in which the work will be completed. Therefore, certain agents
may be used at different Biological Safety Levels depending upon the circumstances. For instance,
human clinical samples from HIV-positive patients may be safely handled at BSL-2. Growth of
HIV in culture should be performed under BSL-3 containment. Biological Safety Levels may be
higher or lower than what is given below for a particular agent depending upon the circumstances of
its use.
The EH&S Biological Safety Officer (BSO) reviews all projects involving recombinant DNA,
infectious disease agents, and agents of concern to livestock and agriculture and will assist you in
the risk assessment process. Once the Institutional Biosafety Committee (IBC) and/or the EH&S
BSO assigns a Biological Safety Level, it must be adhered to unless new information to warrant a
change, in most cases from peer-reviewed literature, is provided. The IBC and/or BSO will review
the literature and make an adjustment, if warranted.
Biological Safety Level 1 (BSL-1)
Agents that are not associated with disease in healthy adult humans, are of minimal potential hazard
to laboratory personnel, and of minimal potential hazard to the environment may be used at BSL-1.
Agents that may be used at BSL-1 include Lactobacillus spp., asporogenic Bacillus subtilis or
Bacillus licheniformis, Escherichia coli-K12 (cloning strains), Baculoviruses, and adeno-associated
virus types 1 through 4 in low concentrations (<109 IP/ml)
Those agents not listed under Biological Safety Levels 2, 3 and 4 are not automatically or implicitly
classified as BSL-1; a risk assessment must be conducted based on the known and potential
properties of the agents and their relationship to agents that are listed.
Biological Safety Level 2 (BSL-2)
Agents to be used at BSL-2 are associated with human disease which is rarely serious and for which
preventive or therapeutic interventions are often available. They are of moderate potential hazard to
laboratory personnel and/or the environment.
Biosafety Manual
37
BSL-2 - Bacterial Agents Including Chlamydia
 Acinetobacter baumannii (formerly Acinetobacter calcoaceticus)
 Actinobacillus
 Actinomyces pyogenes (formerly Corynebacterium pyogenes)
 Aeromonas hydrophila
 Amycolata autotrophica
 Archanobacterium haemolyticum (formerly Corynebacterium haemolyticum)
 Arizona hinshawii - all serotypes
 Bartonella henselae, B. quintana, B. vinsonii
 Bordetella including B. pertussis
 Borrelia recurrentis, B. burgdorferi
 Burkholderia (formerly Pseudomonas species) except those listed under BSL-3
 Campylobacter coli, C. fetus, C. jejuni
 Chlamydia psittaci (non-avian strains), trachomatis, C. pneumoniae
 Clostridium haemolyticum, Cl. histolyticum, Cl. novyi, Cl. septicum, Cl. tetani
 Corynebacterium diphtheriae, C. pseudotuberculosis, C. renale
 Dermatophilus congolensis
 Edwardsiella tarda
 Erysipelothrix rhusiopathiae
 Escherichia coli - all enteropathogenic, enterotoxigenic, enteroinvasive and strains bearing K1
antigen, including E. coli O157:H7
 Haemophilus ducreyi, H. influenzae
 Helicobacter pylori
 Klebsiella
 Legionella including L. pneumophila
 Leptospira interrogans - all serotypes
 Listeria
 Moraxella
 Mycobacterium (except those listed under BSL-3) including M. avium complex, M. asiaticum,
M. bovis BCG vaccine strain, M. chelonei, M. fortuitum, M. kansasii, M. malmoense, M.
marinum, M. paratuberculosis, M. scrofulaceum, M. simiae, M. szulgai, M. ulcerans, M. xenopi
 Neisseria gonorrhoea, N. meningitidis
 Nocardia asteroides, N. brasiliensis, N. otitidiscaviarum, N. transvalensis
 Rhodococcus equi
 Salmonella including S. arizonae, S. cholerasuis, S. enteritidis, S. gallinarum-pullorum, S.
meleagridis, S. paratyphi, A, B, C, S. typhi, S. typhimurium
 Shigella including S. boydii, S. dysenteriae, type 1, S. flexneri, S. sonnei
 Sphaerophorus necrophorus
 Staphylococcus aureus
 Streptobacillus moniliformis
 Streptococcus including S. pneumoniae, S. pyogenes
 Treponema pallidum, T. carateum
 Vibrio cholerae, V. parahemolyticus, V. vulnificus
 Yersinia enterocolitica
38
Biosafety Manual
BSL-2 - Fungal Agents
 Cryptococcus neoformans
 Dactylaria galopava (Ochroconis gallopavum)
 Epidermophyton
 Exophiala (Wangiella) dermatitidis
 Fonsecaea pedrosoi
 Microsporum
 Penicillium marneffei
 Sporothrix schenckii
 Trichophyton
BSL-2 - Parasitic Agents
 Ancylostoma human hookworms including A. duodenale, A. ceylanicum
 Ascaris including Ascaris lumbricoides suum
 Babesia including B. divergens, B. microti
 Brugia filaria worms including B. malayi, B. timori
 Coccidia
 Cryptosporidium including C. parvum
 Cysticercus cellulosae (hydatid cyst, larva of T. solium)
 Entamoeba histolytica
 Enterobius
 Fasciola including F. gigantica, F. hepatica
 Giardia including G. lamblia
 Heterophyes
 Hymenolepis including H. diminuta, H. nana
 Isospora
 Leishmania ethiopia, L. major, L. mexicana, L. peruvania, L. tropica
 Loa loa filaria worms
 Microsporidium
 Necator human hookworms including N. americanus
 Onchoerca filaria worms including, O. volvulus
 Plasmodium including simian species, P. cynomologi, P. falciparum, P. malariae, P. ovale, P.
vivax
 Sarcocystis including S. sui hominis
 Schistosoma including S. haematobium, S. intercalatum, S. japonicum, S. mansoni, S. mekongi
 Strongyloides including S. stercoralis
 Toxocara including T. canis
 Toxoplasma including T. gondii
 Trichinella spiralis
 Trypanosoma including T. brucei brucei, T. brucei gambiense, T. brucei rhodesiense, T.
cruzi
 Wuchereria bancrofti filaria worms
Biosafety Manual
39
BSL-2 - Viruses
Adenoviruses, human - all types except African Swine Fever virus
Arenaviruses
 Lymphocytic choriomeningitis virus (non-neurotropic strains)
 Other viruses as listed by ABSA/CDC
Bunyaviruses
 Bunyamwera virus
 Rift Valley fever virus vaccine strain MP-12 only
 Other viruses as listed by ABSA/CDC
Calciviruses
 Norovirus
 Sapovirus
Coronaviruses except SARS
Flaviviruses (Togaviruses) - Group B Arboviruses
 Yellow fever virus vaccine strain 17D
 Other viruses as listed by ABSA/CDC
Hepatitis A, B, C, D, and E viruses
Herpesviruses - except Herpesvirus simiae (Monkey B virus), BSL-4
 Cytomegalovirus
 Epstein Barr virus
 Herpesvirus ateles
 Herpesvirus saimiri
 Herpes simplex types 1 and 2
 Herpes zoster
 Human herpesvirus types 6 and 7
Marek's disease virus
Murine cytomegalovirus
Pseudorabies virus
Orthomyxoviruses
 Influenza viruses types A, B, and C
 Other tick-borne orthomyxoviruses as listed by ABSA/CDC
Papovaviruses
 All human papilloma viruses
 Bovine papilloma virus
 Polyoma virus
 Shope papilloma virus
 Simian virus 40 (SV40)
40
Biosafety Manual
Paramyxoviruses
 Newcastle disease virus (non-exotic, non-virulent strains)
 Measles virus
 Mumps virus
 Parainfluenza viruses types 1, 2, 3, and 4
 Respiratory syncytial virus
Parvoviruses
 Human parvovirus (B19)
 Canine Parvovirus (CPV)
Picornaviruses
 Coxsackie viruses types A and B
 Echoviruses - all types
 Polioviruses - all types, wild and attenuated
 Rhinoviruses - all types
Poxviruses
 Vaccinia - all types except Monkeypox virus (BSL-3) and restricted poxviruses including
Alastrim, Camelpox, Goatpox, Smallpox, and Whitepox (restricted to the CDC, Atlanta, GA)
Reoviruses - Coltivirus, human Rotavirus, and Orbivirus (Colorado tick fever virus)
Retroviruses
 Bovine leukemia virus
 Clinical samples from HIV-positive patients
 Feline immunodeficiency virus
 Feline leukemia virus
 Feline sarcoma virus
 Gibbon leukemia virus
 Mason-Pfizer monkey virus
 Mouse mammary tumor virus
 Murine leukemia virus
 Murine sarcoma virus
 Rat leukemia virus
NOTE: Containment requirements for research involving rDNA
Experiments involving recombinant DNA lend themselves the application of highly specific
biological barriers. The containment levels required for research involving recombinant DNA
associated with plants or animals is based on classification of experiments in Section III, of the NIH
document “Experiments Covered by the NIH Guidelines” found at
www4.od.nih.gov/oba/rac/guidelines_02/NIH_Guidelines_Apr_02.htm#_SECTION_III._EXPERI
MENTS. Consult this document for containment requirements specific to your research.
NOTE: Murine Retroviral Vectors
Murine retroviral vectors to be used for human transfer experiments (less than 10 liters) that contain
less than 50% of their respective parental viral genome and that have been demonstrated to be free
Biosafety Manual
41
of detectable replication competent retrovirus can be maintained, handled, and administered, under
BL1 containment.
Rhabdoviruses
 Mokola Virus
 Vertebrate and Invertebrate lyssaviruses
Togaviruses (see Alphaviruses and Flaviviruses)
 Rubivirus (rubella)
 Ross River Virus
Biological Safety Level 3 (BSL-3)
Agents to be used at BSL-3 are associated with serious or lethal human disease for which preventive
or therapeutic interventions may be available.
BSL-3 - Bacterial Agents Including Rickettsia
 Bacillus anthracis
 Bartonella
 Brucella including B. abortus, B. canis, B. suis
 Burkholderia (Pseudomonas) mallei, B. pseudomallei
 Clostridium botulinum, Cl. chauvoei
 Coxiella burnetii
 Francisella tularensis (Types A and B)
 Mycobacterium bovis (except BCG strain, BSL-2), M. tuberculosis, M. leprae
 Mycoplasma, except M. mycoides and M. agalactiae which are restricted animal pathogens
 Pasteurella multocida type B -“buffalo” and other virulent strains
 Rickettsia akari, R. australis, R. canada, R. conorii, R. prowazekii, R. rickettsii, R, siberica, R.
tsutsugamushi, R. typhi (R. mooseri)
 Yersinia pestis
BSL-3 - Fungal Agents
 Blastomyces dermatitidis
 Cladosporium bantianum, C. (Xylohypha) trichoides
 Coccidioides immitis (sporulating cultures; contaminated soil)
 Histoplasma capsulatum, H. capsulatum var.. duboisii
 Paracoccidioides braziliensis
BSL-3 - Parasitic Agents
 Echinococcus including E. granulosis, E. multilocularis, E. vogeli
 Leishmania including L. braziliensis, L. donovani
 Naegleria fowleri
 Plasmodium falciparum
 Taenia solium
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Biosafety Manual
BSL-3 - Viruses and Prions
Alphaviruses (Togaviruses) - Group A Arboviruses
 Eastern equine encephalomyelitis virus
 Venezuelan equine encephalomyelitis vaccine strain TC-83
 Western equine encephalomyelitis virus
 Semliki Forest virus
 Venezuelan equine encephalomyelitis virus (except the vaccine strain TC-83 is
 Other viruses as listed by ABSA/CDC
BSL-2)
Arenaviruses
 Lymphocytic choriomeningitis virus (LCM) (neurotropic strains)
 Flexal
Bunyaviruses
 Hantaviruses including Hantaan virus
 Rift Valley fever virus
 Other viruses as listed by ABSA/CDC
Flaviviruses (Togaviruses) - Group B Arboviruses
 Dengue virus serotypes 1, 2, 3, and 4
 Japanese encephalitis virus
 St. Louis encephalitis virus
 Yellow fever virus – wild type
 Other viruses as listed by ABSA/CDC
Poxviruses
 Monkeypox virus
Prions
 Transmissible spongioform encephalopathies (TME) agents, Creutzfeldt-Jacob
and kuru agents (see BMBL for specific containment instruction)
disease
Retroviruses
 Avian leukosis virus
 Avian sarcoma virus
 Human immunodeficiency virus (HIV) types 1 and 2
 Human T cell lymphotropic virus (HTLV) types 1 and 2
 Simian immunodeficiency virus (SIV)
Rhabdoviruses
 Rabies virus
 Vesicular stomatitis virus - laboratory adapted strains ONLY including VSV-Indiana, San Juan,
and Glasgow
Biosafety Manual
43
Biological Safety Level 4 (BSL-4)
Agents to be used at BSL-4 are likely to cause serious or lethal human disease for which preventive
or therapeutic interventions are not usually available.
BSL-4 - Bacterial Agents
None
BSL-4 - Fungal Agents
None
BSL-4 - Parasitic Agents
None
BSL-4 - Viral Agents
Arenaviruses (Togaviruses) - Group A Arboviruses
 Guanarito virus
 Lassa virus
 Junin virus
 Machupo virus
 Sabia virus
Bunyaviruses (Nairovirus)
 Crimean-Congo hemorrhagic fever virus
Filoviruses
 Ebola virus
 Marburg virus
Flaviruses (Togaviruses) - Group B Arboviruses
 Tick-borne encephalitis virus complex including Absetterov, Central European encephalitis,
Hanzalova, Hypr, Kumlinge, Kyasanur Forest disease, Omsk hemorrhagic fever, and Russian
spring-summer encephalitis viruses
Herpesviruses (alpha)
 Herpesvirus simiae (Herpes B or Monkey B virus)
Paramyxiviruses
 Equine morbillivirus
Hemorrhagic fever agents and viruses as yet undefined.
44
Biosafety Manual
2 - Information for Researchers
Project Registration
Northern Arizona University Insitutional Biosafety Committee
Many research projects involve work with potentially hazardous biological agents, known infectious
disease agents, or biological materials regulated by the federal or state government. Granting
agencies require that the university monitor the use of biological hazards, infectious disease agents,
and recombinant DNA in order for them to release funds to investigators. Therefore, we have
developed a registration system to ensure that all biological materials are handled properly and
disposed of appropriately. The EH&S Biological Safety Office administers four registration
programs for research projects.
The NAU Institutional Biosafety Committee (IBC) reviews all registrations. Approval by the NAU
IBC, in writing, is required before ordering or working with any agents. Please contact the EH&S
BSO for more information.
Fillable registration forms are available on the EH&S website at:
http://nau.edu/Research/Compliance/Environmental-Health-and-Safety/Forms/
Biological Agent (BA) Registration
Use of the following materials requires that the Principal Investigator completes and submits the
bio-agent registration document for approval by the EH&S Biological Safety Officer.
Agents characterized at Biosafety Level 1 require registration. These will be expedited and
administratively approved barring any complications.
Agents to be used at Biosafety Level 2 (BSL-2) or Biosafety Level 3 (BSL-3):
1.
All human, animal, or plant pathogens that require BSL-2 or BSL-3 containment and
handling (see previous section: “Agents List”) must be registered. Please note that BSL-4
agents may not be stored or used at NAU.
2.
Unknown human and animal pathogens must be registered. These are considered BSL-2
until identified.
3.
Cell lines or cultures that
1) have been immortalized with a virus (such as EBV or a retrovirus),
2) are known to be tumorigenic in primates (including humans), or
3) are primary human tumor cells.
These are considered BSL-2 (or higher in many cases).
4.
Human blood or other tissues, when used in research, must be registered.
5.
Human blood or other tissue known to be HIV positive or known to contain any human
disease causing agent may require higher containment depending upon the IBC evaluation.
Recombinant DNA (R-DNA) Registration
All R-DNA projects that involve a recombinant organism (this excludes projects that involve DNA
only, i.e. PCR) require registration with the EH&S Biological Safety Office. A subset of R-DNA
Biosafety Manual
45
projects requires review and approval from the Institutional Biosafety Committee (IBC). The NAU
IBC oversees all research projects and issues involving R-DNA at NAU. Use of the following
requires that the principal investigator completes and submits an R-DNA registration document.
1.
All R-DNA projects, including the growth of recombinant bacteria for probe isolation
(plasmid or phage preparations) require registration. Projects must be registered regardless
of where the material came from or who originally constructed it.
2.
Projects that are exempt from the NIH Guidelines must also be registered.
3.
The development of transgenic animals and plants requires registration.
R-DNA projects are performed at BSL-1, BSL-2, BSL-3 or the corresponding levels for whole plant
(BSL-1P, BSL-2P, BSL-3P) or whole animal (BSL-1N, BSL-2N, BSL-3N) work. The EH&S
Biological Safety Officer, in conjunction with the IBC, will make the final determination.
Acute Toxins (AT) Registration
The use and storage of chemicals with a mammalian LD50 of < 100 g/kg require registration. For a
partial list, see the Toxins Table that follows. Acute toxins may only be ordered following written
approval by the BSO or IBC.
Regulated Biological Materials
Agents, such as plant pathogens or exotic microorganisms, that are regulated by federal or state
agencies (CDC, HHS, DOC, USDA/APHIS, EPA, FDA, DPI, etc.) shall be registered with the
EH&S Biological Safety Office by submission of a biological agent registration form. All permits
for transport, transfer, import or export of these regulated agents are the responsibility of the
Principal Investigator. The NAU Responsible Official (RO) will be responsible for all Select Agent
permits. Please allow a minimum of 6 weeks to obtain new federal permits.
Project Amendment
Changes to an existing registration can be done on an amendment unless said changes result in a
dramatic change to the overall project or the containment level.
Typical changes include:



46
Addition or deletion of new personnel
Addition or deletion of new agents (same containment level)
Minor modifications to the protocol or procedures ( final decision to be made by the IBC)
Biosafety Manual
Table 3: Toxin Table
Toxins with a mammalian LD50 of < 100 g/kg must be registered with the EH&S Biological Safety
Office. Therefore, use of the following toxins requires registration. If a toxin is not on the list, it
still may require registration, depending upon the LD50. For more information, please contact the
EH&S Biological Safety Office.
Toxicity
LD50 (g/kg)*
Abrin
Aerolysin
Botulinin toxin A
Botulinin toxin B
Botulinin toxin C1
Botulinin toxin C2
Botulinin toxin D
Botulinin toxin E
Botulinin toxin F
-bungarotoxin
Caeruleotoxin
Cereolysin
Cholera toxin
Clostridium difficile enterotoxin A
Clostridium difficile cytotoxin B
Clostridium perfringens lecithinase
Clostridium perfringens kappa toxin
Clostridium perfringens perfringolysin O
Clostridium perfringens enterotoxin
Clostridium perfringens beta toxin
Clostridium perfringens delta toxin
Clostridium perfringens epsilon toxin
Conotoxin
Crotoxin
Diphtheria toxin
Listeriolysin
Leucocidin
Modeccin
Nematocyst toxins
Notexin
Pertussis toxin
Biosafety Manual
0.7
7.0
0.0012
0.0012
0.0011
0.0012
0.0004
0.0011
0.0025
14.0
53
40-80
250
0.5
220
3
1500
13-16
81
400
5
0.1
12-30
82
0.1
3-12
50
1-10
33-70
25
15
47
Toxicity
LD50 (g/kg)*
Pneumolysin
Pseudomonas aeruginosa toxin A
Ricin
Saxitoxin
Shiga toxin
Shigella dysenteriae neurotoxin
Streptolysin O
Staphylococcus enterotoxin B
Staphylococcus enterotoxin F
Streptolysin S
Taipoxin
Tetanus toxin
Tetrodotoxin
Viscumin
Volkensin
Yersinia pestis murine toxin
1.5
3
2.7
8
0.250
1.3
8
25
2-10
25
2
0.001
8
2.4-80
1.4
10
*Please note that the LD50 values are from a number of sources (see below). For specifics on route
of application (i.v., i.p., s.c.), animal used, and variations on the listed toxins, please go to the
references listed below.
Reference:
1.
Gill, D. Michael; 1982; Bacterial toxins: a table of lethal amounts; Microbiological Reviews;
46: 86-94
2.
Stirpe, F.; Luigi Barbieri; Maria Giulia Battelli, Marco Soria and Douglas A. Lappi; 1992;
Ribosome-inactivating proteins from plants: present status and future prospects;
Biotechnology; 10: 405-412
3.
Registry of toxic effects of chemical substances (RTECS): comprehensive guide to the
RTECS. 1997. Doris V. Sweet, ed., U.S. Dept of Health and Human Services, Public Health
Service, Centers for Disease Control and Prevention, National Institute for Occupational
Safety and Health; Cincinnati, Ohio
Select Agents
The following lists of agents and toxins are classified by the Federal government as Select Agents.
Any possession, use, transfer or shipment of these materials is strictly controlled by regulation.
Researchers considering work with any of these materials must first contact the NAU Responsible
Official for the approvals, permits, clearances and other necessary paperwork. Be aware that
government clearance can take as much as 6 months to complete in advance of any project.
Failure to comply with these Federal Regulations is punishable by both fines and imprisonment.
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Biosafety Manual
HHS and USDA Select Agents and Toxins
7CFR Part 331, 9 CFR Part 121, and 42 CFR Part 73
HHS SELECT AGENTS AND TOXINS
Abrin
Botulinum neurotoxins*
Botulinum neurotoxin producing species
of Clostridium*
Conotoxins (Short, paralytic alpha conotoxins
containing the following amino acid sequence
X1CCX2PACGX3X4X5X6CX7)1
Coxiella burnetii
Crimean-Congo haemorrhagic fever virus
Diacetoxyscirpenol
Eastern Equine Encephalitis virus3
Ebola virus*
Francisella tularensis*
Lassa fever virus
Lujo virus
Marburg virus*
Monkeypox virus3
Reconstructed replication competent forms of the
1918 pandemic influenza virus containing any
portion of the coding regions of all eight gene
segments (Reconstructed 1918 Influenza virus)
Ricin
Rickettsia prowazekii
SARS-associated coronavirus (SARS-CoV)
Saxitoxin
South American Haemorrhagic Fever viruses:
Chapare
Guanarito
Junin
Machupo
Sabia
Staphylococcal enterotoxins A,B,C,D,E subtypes
T-2 toxin
Tetrodotoxin
Tick-borne encephalitis complex (flavi) viruses:
Far Eastern subtype
Siberian subtype
Kyasanur Forest disease virus
Omsk hemorrhagic fever virus
Variola major virus (Smallpox virus)*
Variola minor virus (Alastrim)*
Yersinia pestis*
OVERLAP SELECT AGENTS AND TOXINS
Bacillus anthracis*
Bacillus anthracis Pasteur strain
Brucella abortus
Brucella melitensis
Brucella suis
Burkholderia mallei*
Burkholderia pseudomallei*
Hendra virus
Nipah virus
Rift Valley fever virus
Venezuelan equine encephalitis virus3
USDA SELECT AGENTS AND TOXINS
African horse sickness virus
African swine fever virus
Avian influenza virus3
Classical swine fever virus
Foot-and-mouth disease virus*
Goat pox virus
Lumpy skin disease virus
Mycoplasma capricolum3
Mycoplasma mycoides3
Newcastle disease virus2,3
Peste des petits ruminants virus
Rinderpest virus*
Sheep pox virus
Swine vesicular disease virus
USDA PLANT PROTECTION AND QUARANTINE (PPQ)
SELECT AGENTS AND TOXINS
Peronosclerospora philippinensis
(Peronosclerospora sacchari)
Phoma glycinicola (formerly Pyrenochaeta glycines)
Ralstonia solanacearum
Rathayibacter toxicus
Sclerophthora rayssiae
Synchytrium endobioticum
Xanthomonas oryzae
*Denotes Tier 1 Agent
Biosafety Manual
49
1
C = Cysteine residues are all present as disulfides, with the 1st and 3rd Cysteine, and the 2nd and 4th
Cysteine forming specific disulfide bridges; The consensus sequence includes known toxins α-MI and α-GI
(shown above) as well as α-GIA, Ac1.1a, α-CnIA, α-CnIB; X1 = any amino acid(s) or Des-X; X2 = Asparagine
or Histidine; P = Proline; A = Alanine; G = Glycine; X3 = Arginine or Lysine; X4 = Asparagine, Histidine,
Lysine, Arginine, Tyrosine, Phenylalanine or Tryptophan; X5 = Tyrosine, Phenylalanine, or Tryptophan; X6
= Serine, Threonine, Glutamate, Aspartate, Glutamine, or Asparagine; X7 = Any amino acid(s) or Des X
and; “Des X” = “an amino acid does not have to be present at this position.” For example if a peptide
sequence were XCCHPA then the related peptide CCHPA would be designated as Des-X.
2
A virulent Newcastle disease virus (avian paramyxovirus serotype 1) has an intracerebral pathogenicity
index in day-old chicks (Gallus gallus) of 0.7 or greater or has an amino acid sequence at the fusion (F)
protein cleavage site that is consistent with virulent strains of Newcastle disease virus. A failure to detect
a cleavage site that is consistent with virulent strains does not confirm the absence of a virulent virus.
3
Select agents that meet any of the following criteria are excluded from the requirements of this part:
Any low pathogenic strains of avian influenza virus, South American genotype of eastern equine
encephalitis virus , west African clade of Monkeypox viruses, any strain of Newcastle disease virus which
does not meet the criteria for virulent Newcastle disease virus, all subspecies Mycoplasma capricolum
except subspecies capripneumoniae (contagious caprine pleuropneumonia), all subspecies Mycoplasma
mycoides except subspecies mycoides small colony (Mmm SC) (contagious bovine pleuropneumonia), and
any subtypes of Venezuelan equine encephalitis virus except for Subtypes IAB or IC, provided that the
individual or entity can verify that the agent is within the exclusion category. 9/10/13
Minors in Research Laboratories or Animal Facilities
Unless enrolled as a Northern Arizona University student, minors are not allowed to work or
conduct research in Northern Arizona University research laboratories, greenhouses or animal
facilities except as identified specifically below. In addition, minors are prohibited from operating
farm machinery or state vehicles and from working in machine shops.
1. All Minors are prohibited from working or conducting research in the following:
a. Any laboratory or facility designated as BSL-3, ABSL-3 or higher for recombinant or
infectious organisms.
b. Any laboratory where select agents or explosives are used or stored.
c. Any Animal Care Services (ACS) housing or procedure area/lab/facility.
2. Minors are prohibited from working with any of the following materials.
a. Radioactive materials or radiation (X-rays)
b. Acute Toxins
3. Minors are allowed to work or conduct research in laboratories (not listed in #1 or #2 above)
if the following requirements are met in full:
a. Northern Arizona University EH&S Biosafety Office Policy titled; Minors In
Research Laboratories Or Animal Facilities
(www4.nau.edu/ovp/regulatorycompliance/safety/MinorsResearchPolicy.asp.)
has been read and understood. The Potential Hazards Information & Signature
Sheet (www4.nau.edu/ovp/regulatorycompliance/safety/Potentialhazardssheet.asp.)
has been reviewed, signed by the parent/legal guardian and returned to the EH&S
Biosafety Office by mail or by fax.
b. A Minor’s Research Proposal Registration Form
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Biosafety Manual
c.
d.
e.
f.
(www4.nau.edu/ovp/regulatorycompliance/biosafety/forms/minorsregistrationform.
doc.) is submitted to and approved by Northern Arizona University Institutional
Biosafety Committee which includes:
i. A description of the project and all materials
ii. A parental consent form
iii. A sponsor consent form
Hazard specific safety training is completed by the Principal Investigator/Sponsor
with the minor as approved by EH&S BSO.
Personal protective equipment, specific to the hazard, is provided to the minor with
instructions for use and disposal.
The minor is supervised at all times while in the laboratory and never left alone.
Hours of work comply with Federal Regulation 29 CFR 570.35.
The laboratory is in full compliance with all applicable Northern Arizona University
safety programs and regulations.
Responsibilities of the Principle Investigator during the
Conduct of the Research
The Principal Investigator shall:
1.
2.
3.
4.
5.
6.
Supervise the safety performance of the laboratory staff to ensure that the required safety
practices and techniques are employed.
Report any significant incident, violation of the NIH Guidelines, or any significant,
research-related accidents and illnesses immediately by contacting the Biosafety Officer.
Examples of incidents and violations include:
a. Overt exposures (exposures that result in direct personnel exposure to biohazards
such as injection, spills, splashes or aerosol inhalation)
b. Potential exposures (exposures that have a high risk of exposing personnel to
biohazards such as spills, containment failure while working with the agent or
equipment failure that may produce aerosols)
c. Any exposure (overt or potential) in a BSL-3 laboratory
d. Overt exposure in BSL1 or BSL-2 laboratories
e. Any illness that may be caused by the agents used in the laboratory
f. Incidents involving the improper disposal of biohazards.
Ensure the integrity of the physical containment (e.g., biological safety cabinets) and the
biological containment (e.g., purity and genotypic and phenotypic characteristics) and
correct procedures or conditions that might result in release of or exposure to biohazards.
Limit or restrict access to the laboratory when work with biohazards is in progress; this
includes making a determination of who may be at increased risk.
Establish policies and procedures to limit access exclusively to those individuals who have
been advised of the potential hazards and meet specific entry requirements.
Ensure that laboratory personnel are offered, at no cost, appropriate immunizations or tests
for the infectious agents handled or potentially present in the laboratory (e.g., hepatitis B
vaccine, tuberculosis skin testing).
Biosafety Manual
51
Reportable Incidents and Violations
Incidents/problems involving biohazards must be immediately reported to the Biosafety Officer.
Examples of reportable significant incidents include but are not limited to:
1. Any overt exposure, such as a needle stick, splash, and contamination due to equipment failure.
2. Any potential exposure in a BSL-3 facility.
A significant event may also occur from a containment breach, which may be subsequently
determined to pose either an overt or potential exposure to individuals or the environment. It should
be noted that waste from recombinant or synthetic nucleic acid research is also considered
biohazardous and incidents involving improper disposal of recombinant or synthetic nucleic acids
must also be reported. Questions regarding reportable incidents should be directed to the Biosafety
Officer.
Failure by research personnel to follow federal and institutional regulations, guidelines, policies
and/or procedures may also require reporting to the appropriate institutional, local, state and/or
federal agencies. Violations may include but are not limited to conduct of new or ongoing research
without appropriate federal or institutional registration, review, approval or oversight.
Institutional Reporting Responsibilities
The Institutional Biosafety Committee is required, by the NIH Guidelines, to report to the
appropriate University official and to the NIH/OBA within thirty days any significant incidents,
violations of the NIH Guidelines, or any significant findings of research-related accidents and
illnesses. The IBC will be responsible to determine what actions, if any, are necessary. For example,
the IBC may determine the need to make changes to the frequency of laboratory inspections or
biosafety containment level of the research, based on results of the incident.
Other IBC reporting requirements (to OBA and other agencies) include but are not limited to:
1. Research involving biohazards conducted without prior IBC approval
2. Lax security, unsafe procedures used in a laboratory setting, improper disposal of
recombinant or synthetic nucleic acid waste
3. Changes to research risk that have been initiated without prior approval by IBC
Some incidents must be reported to OBA on an expedited basis. Spills or accidents in BSL-2
laboratories involving recombinant or synthetic nucleic acids that result in an overt exposure must
be immediately reported to OBA. In addition, spills or accidents involving recombinant or synthetic
nucleic acids occurring in high containment (BSL-3 or higher) laboratories resulting in an overt or
potential exposure must be immediately reported to OBA. The IBC working through the IBC Chair
and the BSO will report to the Institutional Official, who, in turn will oversee the report to OBA,
any of the above-described incidents.
Institutional violations that will also be reported to the appropriate College or department head may
include but are not limited to:
1. Lapses in disclosure approval
2. Failure to comply with institutional and federal regulations, guidelines, and policies
3. Unsafe work practices
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Biosafety Manual
Biological Waste Disposal Policy
This policy is intended to provide guidance and insure compliance with the NIH/CDC guidelines,
the State of Arizona Administrative Code, and restrictions of the Coconino County Landfill.
Categories
1) Infectious/potentially infectious/R-DNA
a) human pathogens
b) animal pathogens
c) plant pathogens
d) recombinant DNA
e) human and primate blood, blood products and other body fluids
f) human and primate tissue
g) any material containing or contaminated with any of the above (test tubes, needles*,
syringes, tubing, culture dishes, flasks, etc.)
This waste must be inactivated prior to disposal. The preferred method is steam sterilization
(autoclaving), although chemical inactivation or incineration may be appropriate in some cases. Per
NAU policy, even autoclaved or otherwise inactivated wastes must be disposed of as biohazardous
waste. Storage of non-inactivated waste is restricted to within the generating laboratory. The
material may not be stored longer than 24 hours prior to inactivation.
2) Non-infectious waste
This category includes waste that is not contaminated with any of the biological materials listed in
category 1. It includes solid waste and sharps generated in clinical or laboratory settings. Sterile or
unopened biomedical materials that require disposal are also considered biological waste, and must
be treated as such.
IV packs
needles*
syringes
scalpels*
broken glass and plastic ware**
test tubes
razor blades*
culture dishes
flasks
pipettes
Petri dishes
tissue culture flasks
This material does not require sterilization prior to disposal.
*must be packaged in plastic sharps boxes.
**must be within a box or other puncture proof container before adding to waste.
3) Mixed radioactive/biohazardous waste
The biohazardous component of mixed radioactive/biohazardous waste shall be inactivated prior to
its release to Radiation Safety for disposal as radioactive waste. Steam-sterilization or chemical
inactivation shall be employed as above. Although some radioactive materials can be autoclaved
safely, please check with the EH&S Radiation Safety Officer regarding the best method to employ
with any given radionuclide.
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53
4) Mixed chemical/biohazardous waste
The biohazardous component of mixed chemical/biohazardous waste shall be inactivated prior to its
release for chemical disposal. Precautions should be taken to prevent the generation and release of
toxic chemicals during the inactivation process. In general, autoclaving is not recommended
because flammable or reactive compounds should not be autoclaved due to the explosion hazard.
Please check with the NAU Chemical Hygiene Officer for guidance regarding particular chemicals.
5) Animal carcasses and materials
As there are no tissue digesters or incinerators present on the NAU
Campus, all animal materials must be packaged, shrouded and delivered
to the Vivarium Manager. No animal bodies or material shall be disposed
of as regular trash or through the biomedical waste receptacle.
6) Human remains
Please contact the State Anatomical Review Board for information
regarding the final disposition of human remains and body parts.
Packaging
1) Biohazard bags
These are used for the initial collection of certain biological wastes. All biohazard bags must meet
impact resistance (165 grams), tearing resistance (480 grams), and heavy metal concentration (<100
PPM total of lead, mercury, chromium and cadmium) requirements. Written documentation (a test
report) from the manufacturer regarding these requirements must be on file. Individual departments
are responsible for obtaining the appropriate biohazard bags. These bags must be placed in bulk
biohazard containers as described within this section.
2) Sharps
Needles, scalpels and razor blades must be containerized in red plastic sharps
containers. NAU departments are responsible for obtaining their own sharps
containers. All other sharps (broken glass and plastic ware, pipettes, etc.) must be
containerized in puncture-resistant cardboard boxes (see #3, below).
3) Containers
All biological waste must be containerized in rigid, leak proof, puncture resistant boxes or plastic
tubs/barrels as the terminal receptacle. NAU Departments are responsible for obtaining the
appropriate containers from the EH&S Hazardous Waste Coordinator.
Labeling
All packages containing biological waste shall be labeled with indelible ink marker (i.e., Sharpie®)
as follows:
1) Date
Biohazard bags shall be labeled with the date they were put into use. Please note that biohazard
bags must be labeled even though they will be placed inside a secondary container for final disposal.
Sharps containers shall be labeled with the date the container is full.
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Biosafety Manual
2) Name/Location/Phone Number
Generator’s (principal investigator’s) name, lab location (room number) and phone number will be
clearly printed on each container.
3) Biohazard sign
Only manufactured containers with the preprinted universal biohazard symbol and the words
“biomedical”, “biohazardous”, or “infectious” shall be used.
Transport
The transport of biohazardous waste outside of the laboratory (i.e., to an autoclave or incinerator)
must be in a closed, leak-proof container that is labeled “biohazard”. Only trained personnel may
transport biomedical waste. Labeling may be accomplished by use of a red biohazard bag with the
universal biohazard symbol. Only biomedical bags and red plastic sharps containers may be used to
transport biological waste to the biomedical waste receptacle. Waste receptacle personnel are
instructed not to accept any other type of containers.
Transportation of red-bagged waste must be in closed, leak-proof containers, properly labeled as
“Biohazards.” Movement of regulated/biological waste through public corridors, along carpeted
hallways, and on public elevators should be avoided whenever possible. Any leakage/spills from
these containers must be immediately reported to the NAU Biosafety Office. Signs must be
displayed to prevent tracking of the spills to other areas.
Training
All employees who handle biological waste shall be trained annually regarding its proper handling.
All new employees shall be trained before they are allowed to handle biological waste.
Training may be accomplished through the NAU Bloodborne Pathogen Training Program,
informally in the lab setting, or through formal training programs set up by individual departments
or divisions. For assistance, please call the NAU Biological Safety Officer or Hazardous Waste
Coordinator.
According to Arizona Statute (Ch. 64E-16 F.A.C.), records of the training session shall be
maintained for each employee, along with an outline of the training program. Training records shall
be retained for a period of three (3) years.
Biological Waste Disposal Containers
The following waste disposal containers for biohazardous or biomedical waste are to be used for
teaching and research purposes at Northern Arizona University:
ALL BIOHAZARDOUS WASTE MUST BE TREATED BEFORE DISPOSAL.
Biosafety Manual
55
Bulk Biohazard Receptacles
The EH&S currently provides departments who generate biohazardous and biomedical waste with
large receptacles for the disposal of full biohazard bags and sharps containers. Individual
departments can contact the EH&S Hazardous Waste Coordinator for pickup when those containers
are full.
PLEASE NOTE: PER NAU POLICY, EVEN AUTOCLAVED WASTES MUST BE
TREATED AS BIOHAZARDOUS WASTES.
Disposal of biomedical waste in the regular trash or dumpster is prohibited. The Coconino County
Landfill does not accept any items that resemble “biomedical devices” and will reject the entire
load. In some cases, the individual principal investigator has been made responsible for the costs
incurred for sorting and disposal of improperly disposed waste.
Sharps Boxes
NAU is subject to the requirements of Arizona Administrative Code Rule
18-13-1419, Medical Sharps. This regulation requires that a generator
who treats bio-hazardous medical waste on site shall place medical sharps
in a sharps container after rendering them incapable of creating a stick
hazard by using an encapsulation agent or any other process that prevents
a stick hazard.
Red plastic sharps boxes are used for disposal of needles, razor blades,
scalpels, and small Pasteur pipettes. Sharps boxes that contain infectious
materials must be inactivated by autoclaving the closed box. These boxes
shall be labeled (with indelible ink marker as above) with the principal
investigator’s name, phone number, date the sharps box is full, and lab location (room number).
Only trained staff/research staff shall handle the full sharps boxes. Full sharps containers can be
disposed of in the bulk biohazard receptacles described above.
Biohazard Bags
Red plastic biohazard bags shall be used for collection of biohazardous tissue culture items, petri
dishes, and other non-sharp items. These biohazard bags shall be immediately labeled with an
indelible ink marker (e.g. Sharpie) when the bag is first put into use. Labels shall include the
principal investigator’s name, phone number, the date the first biohazard item is placed into the bag,
and lab location (room number). The biohazardous waste items must be inactivated by autoclaving
in biohazard autoclave bags or treatment with bleach within 30 days of accumulation (Ch. 64E-16
F.A.C.). After treatment, the bags shall be tightly sealed and placed in the bulk containers described
above. They may be transported to the biomedical waste receptacle by trained laboratory personnel
or by trained staff. Biohazard bags should never be handled by non-research staff or placed in the
hallway. Double bagging may be required to prevent leakage, however, absorbent material must be
added to prevent the presence of any liquid or fluid. Full, untreated biohazard bags shall be stored
only in the lab where accumulation occurred. Full biohazard bags must be treated within 24 hours.
As previously noted, even autoclaved wastes must be disposed of as biohazardous waste.
56
Biosafety Manual
Autoclave Use and Testing
Rules governing the use and testing of autoclaves are based on Arizona Administrative Code, Rule
18-13-1405, Biohazardous Medical Waste Treated on Site. The following summarize the
requirements set by this administrative rule.
A generator who autoclaves bio-hazardous medical waste on site shall:
1)
Further process by grinding, shredding, or any other process, any recognizable
animals and human tissue, organs, or body parts, to render such wasted nonrecognizable and ensure effective treatment.
Operate the autoclave at the manufacturer's specifications appropriate for the quantity
and density of the load.
Keep records of operational performance levels for six months after each treatment
cycle. Operational performance level recordkeeping includes all of the following:
a. Duration of time for each treatment cycle.
b. The temperature and pressure maintained in the treatment unit during each cycle.
c. The method used to determine treatment parameters in the manufacturer's
specifications.
d. The method in manufacturer's specifications used to confirm microbial
inactivation and the test results.
e. Any other operating parameters in the manufacturer's specifications for each
treatment cycle.
Keep records of equipment maintenance for the duration of equipment use that
include the date and result of all equipment calibration and maintenance.
2)
3.)
4)
A generator shall also do the following:
1)
2)
3)
4)
Package the treated medical waste according to the waste collection agency's
requirements;
Attach to the package or container a label, placard, or tag with the following
words: "This medical waste has been treated as required by the Arizona
Department of Environmental Quality standards" before placing the treated medical
waste out for collection as a general solid waste. The generator shall ensure that the
treated medical waste meets the standards of R18-13-1415.
Upon request of the solid waste collection agency or municipal solid waste landfill,
provide a certification that the treated medical waste meets the standards of R18-131415.
Make treatment records available for Departmental inspection upon request.
A generator of medical sharps shall handle medical sharps as prescribed in R18-13-1419.
A generator of chemotherapy waste, cultures and stocks, or animal waste shall handle that waste as
prescribed in R18-13-1420.
Autoclave Testing
Autoclaves shall be tested before being placed into service and then periodically for effectiveness.
1)
Periodicity
a)
Every 40 hours of use
Required for autoclaves that are used to inactivate human or non-human primate
blood, tissues, clinical samples, or human pathogens.
Biosafety Manual
57
b)
2)
Annually
Required for autoclaves that are used to inactivate other material.
Method
A commercially available test indicator kit that uses bacterial spores (Bacillus
stearothermophilus) is the approved method of testing autoclave efficiency. Most spore vial
test kits require 56 to 60 C incubation of the autoclaved test vial along with a nonautoclaved control vial. Incubation causes surviving spores to grow.
a)
New autoclaves
Before placing an autoclave into service, a test load approximating the weight and
density of the type of waste generated shall be autoclaved with test spore vials. The
spore vials should be placed at the bottom, top, front, rear, and center of the
autoclave chamber. This can be achieved by either:
-placing vials at those positions within one large test load, OR
-making several smaller test packs with 1 vial at the center of each
and placing the packs at those locations within the chamber.
The appropriate parameters for sterilization including temperature, pressure, and treatment
time shall be determined in this way.
b)
3)
Autoclaves already in use
For periodic testing, place a spore vial in the very center of a test load prior to
autoclaving.
Storage Information
Please read the product information sheet for appropriate storage information. Spore vials
should not be frozen. Each batch of vials has an expiration date. Vials should not be used
after their expiration date.
Record-Keeping
The following records regarding autoclave use must be kept:
1)
On-site maintenance records
2)
Autoclave use log – must be available near the autoclave.
Each load of material inactivated shall be logged as follows:
a)
Date, time, and operator's name
b)
Contact Information: Lab, room number, phone number
c)
Is this biohazardous material?
d)
Confirmation of sterilization:
i)
Record the temperature, pressure, and length of time the load is sterilized.
Please note that temperature sensitive autoclave tape is not sufficient to
indicate that the load reached sterilization conditions because the tape will
change color at lower temperatures.
ii)
Save the autoclave printout, if the autoclave has a working printer.
Autoclave Operating Procedures
A written sterilization procedure shall be in place for each workplace. This shall include the
following:
1)
Parameters
58
Biosafety Manual
2)
3)
Appropriate parameters for sterilization shall be determined from the testing with spore
vials. The time it takes to sterilize a load will change depending upon the load density and
the sterilization cycle one chooses. Tests have been performed which imitate these various
situations. Please follow the established guidelines.
Protocol
Identification of standard treatment containers and proper load placement shall be made.
Cleaning
The autoclave and work areas shall be cleaned after every use and the work area shall be
disinfected, as needed.
Autoclave Operation and Safety Training
Autoclave training is available from the EH&S Biological Safety Office upon request. The training
is geared toward the research staff and goes over proper use of autoclaves and how they may be
maintained and used properly. Safety training is also given.
Biosafety Manual
59
Autoclave Guidelines
AUTOCLAVE GUIDELINES
STERILIZATION TIMES (Drying Time Not Included)
121 C / 250 F and 15 p.s.i.
BIO WASTE IN AUTOCLAVE BAGS, LOOSELY TIED
MULTIPLE BAGS
--- 100 MINUTES OR LONGER
SINGLE BAG (FULL) --- 90 MINUTES
PARTIAL BAG
--- 60 MINUTES
DRY GOODS
GLASSWARE, EMPTY, INVERTED
--- 15 MINUTES
INSTRUMENTS, WRAPPED
--- 30 MINUTES
UTENSILS, WRAPPED
--- 30 MINUTES
LIQUIDS (Bottles with vented caps ½ FULL)
75ml
250ml
500ml
1000ml
1500ml
2000ml
-------------
25 MINUTES
30 MINUTES
40 MINUTES
45 MINUTES
50 MINUTES
55 MINUTES
The above times should be used as a guide in determining the length of time items should be
autoclaved in order to achieve sterilization.
60
Biosafety Manual
Disinfectants
“Disinfectant” refers to an agent that is applied to treat (usually) inanimate objects to render them
free of pathogenic or infectious microorganisms. In contrast, the term, “sterilant”, refers to an agent
that renders items free of all microorganisms. The two are not the same and should not be confused.
Disinfectants are used in laboratory and chemical settings to 1) treat a surface or an item before or
after routine use, or 2) to treat a surface or an item after a spill or “contaminating event”.
Because disinfectants are antimicrobial, they may, by their nature, also have a toxic effect to the
user. Therefore, Material Safety Data Sheets and other manufacturer’s product information should
be available and thoroughly reviewed before using these products.
There are many different types and formulations of disinfectants. The researcher or clinician should
ensure that the proper product, one that is effective against the specific microorganism being
studied, is used.
The FDA regulates those products that are marketed as sterilants or sanitizing agents on medical
devices. They have published a list of products currently on the market that are labeled as sterilants.
(http://www.epa.gov/spdpublc/snap/sterilants/index.html)
The EPA regulates moderate and low level disinfectants as “chemical germicides” under their
pesticide regulations. They have published a list of 1) those products that are effective against
Mycobacterium spp. (tubercle bacilli), and 2) those products that are effective against HIV-1
(human immunodeficiency virus).
Please contact the Biological Safety Office for information about any of these lists or for a list of
manufacturers. Be aware that most disinfectants assume pre-cleaning to remove gross
organic/protein prior to use.
Whenever a disinfectant or sterilant is used, proper safety precautions must be followed.
Appropriate clothing (gloves, safety goggles, aprons) must be worn. In addition, these compounds
must be used in well-ventilated areas.
Following is a discussion of general categories of disinfectants. Please note that there are several
different products and different formulations in each category.
Liquids
Alcohols
The most commonly used alcohols, ethanol and isopropanol, are most effective at concentrations of
70% in water. Both higher and lower concentrations are less effective. Alcohols are active against
vegetative bacteria, fungi, and lipid viruses but not against spores. They are only moderately
effective against nonlipid viruses. Alcohols are difficult to use as contact disinfectants because they
evaporate rapidly and do not penetrate organic matter well. When using alcohols, it is best to clean
an object, then submerge it in alcohol for the appropriate time. Alcohols are often used in concert
with other disinfectants such as formaldehyde (but see toxicity warning below) or chlorine (2000
ppm chlorine in alcohol). Alcohol is NOT a registered tuberculocidal or HIV listed disinfectant.
Biosafety Manual
61
Chlorine compounds
The most commonly used and generally effective disinfectant is sodium hypochlorite (common
household bleach). It is a strong oxidizing agent and therefore can be corrosive to metal. A 1:50
dilution, supplying 1000 ppm available chlorine, of the common household product (e.g. chlorox) is
very effective as a general laboratory disinfectant and a 1:10 dilution supplying 5000 ppm available
chlorine is effective against spills involving blood or other organic material. Please note that the
presence of high concentrations of protein can inactivate the action of chlorine products. Dilute
hypochlorite solution must be prepared daily to be maximally effective. There are more
concentrated sodium hypochlorite solutions available for industrial use, so please read the product
information carefully to determine the proper dilution.
Table 4: Dilutions of Household Bleach
Volume
Of Bleach
Undiluted
1
1
Volume
of Water
0
9
99
Dilution
Ratio
1:1
1:10
1:100
Sodium
Available
Hypochlorite % Chlorine
(mg/L)
5.25
50,000
0.5
5,000
0.05
500
Formaldehyde
Formaldehyde is a gas that is available either dissolved in water and methanol as a 37%
formaldehyde solution, known as formalin, or as a solid, paraformaldehyde, that may be melted to
release the gas. Formaldehyde gas is very active against a variety of microorganisms and is used for
space decontamination and to decontaminate biological safety cabinets. Formaldehyde dissolved in
water is active at 1-8% solutions and can be used to decontaminate hard surfaces. However,
because formaldehyde is very irritating at low concentrations (0.1 to 5 ppm) and a known
carcinogen, its use as a hard surface disinfectant is limited to situations in which it is particularly
needed. Due to its toxic effects, there are no EPA-registered disinfectants that contain
formaldehyde.
Glutaraldehyde
Glutaraldehyde is usually supplied as a 20% solution and requires activation by the addition of an
alkaline agent prior to use. The activated product may be kept for about two weeks and should be
discarded when turbid. Glutaraldehyde is active against almost all microorganisms, but is toxic,
irritating, and mutagenic and should be used only when necessary. Please follow the manufacturer’s
guidance when using glutaraldehyde-based products because there are many different formulations
that have been designed for specific uses.
Hydrogen peroxide
Hydrogen peroxide is usually available as a 30% solution. It may be diluted 1:5 for use as a
disinfectant. It is active against a wide array of microorganisms. However, it is an oxidizing agent
and should not be used on aluminum, copper, zinc, or brass. Hydrogen peroxide is unstable at high
temperatures and in light.
Iodine and Iodophors
Iodine and iodophors, compounds in which the iodine is combined with a solubilizing or carrier
agent, are general, all-purpose disinfectants with an action similar to that of chlorine products. The
appropriate concentration for iodine-containing products is 75 ppm available iodine for disinfecting
work surfaces. Concentrations may be much higher for other purposes. Like chlorine compounds,
62
Biosafety Manual
the effectiveness of iodine compounds may be diminished in the presence of protein/organic
material. Iodophor compounds that are used for antisepsis (germicide applied to tissue or skin) are
not appropriate for use as hard surface disinfectants and vice versa. Please read the product material
for appropriate dilutions and applications.
Phenol and phenolic compounds
Phenolic compounds are active at 0.2 to 3% concentrations against all forms of vegetative
microorganisms but not against spores. They have only limited effectiveness against nonlipid
viruses. There are many common disinfectants based on phenol and they should be used according
to the manufacturer’s recommendations. Phenol is a hazardous chemical and requires hazard
assessment as well as proper PPE selection and use.
Quatenary ammonium compounds
Compounds in this class are active at concentrations of 0.1 - 2%. They are active against vegetative
bacteria, lipid viruses, but not against bacterial spores, non-lipid viruses, or tubercle bacilli. These
compounds should be used only when a low-level disinfectant is required.
Gases
Ethylene Oxide (Not currently allowed at NAU)
Inactivates micro-organisms, cellular disruption
Temp 50-60 C, Humidity 30-40%
Conc 400-500 mg/liter
Time 2 hrs at 60 C or 24 hrs at 20 C
Aeration required after cycle for 8 hrs
Biological indicators needed to confirm kill
Suspected carcinogen with explosive properties
Vapor Phase Hydrogen Peroxide
Temp 4-60 C,> temp results in > activity
Conc 30%, less than 10 mg/liter
Non-toxic end products of water and oxygen
Limited to surfaces, no penetration
Corrosive to some materials
Degrades natural rubber and nylon
Chlorine Dioxide gas
Dilute chlorine gas and sodium chlorite, less than 25 mg/liter
Temp 25-30 C, prehumidification required
Limited to surfaces, no penetration
Corrosive to some materials
Mucous membrane irritant
Ozone
Oxidizing properties, inhibits bacterial growth, reacts with amino acids,
RNA/DNA
Temp 25 C
Conc 2-5 mg/liter
Systems convert oxygen to ozone
Biosafety Manual
63
Limited penetration
Formaldehyde Gas (from heating paraformaldehyde)
Temp 20-22 C, humidity 60-85%
Conc 0.3 gm/cu ft of volume
Time 6-8 hours
Toxic irritant and suspected carcinogen
Limited penetration, primarily surface action
Requires aeration and time for formaldehyde to off-gas, usually 8 hours
Irradiation
Ultraviolet, UV radiation
Virucidal action correlates with shorter wavelengths of the UV spectrum,
250-260 nm.
Mechanism of UV radiation injury attributed to absorption by and resultant damage to
nucleic acids
Due to low energy, the power of penetration is poor.
Dust and thin layers of proteins on surfaces reduce the virucidal activity
Do not rely on Biosafety Cabinet UV lights to disinfect surfaces.
Ionizing radiation
Penetrates products and micro-organisms
Releases high energy, disrupts cells, DNA & RNA and results in kill
Damage done to cell membrane and other cellular structures
Cobalt 60 and Cesium 137
Electron Beam
Limited penetration
Accelerator generates high energy electrons
Microwaves
Thermal and non-thermal activity
Still under study for the most part
Disinfectants Bibliography
The following materials were consulted or used in this chapter:
Disinfection, Sterilization, and Preservation. Fourth edition. 1991. Seymour S. Block ed., Lea & Febiger, Philidelphia
Laboratory Biosafety Manual. Second edition. 1993. World Health Organization, Geneva.
Laboratory Safety: Principles and practices. Second edition. 1995. Diane O. Fleming et al. eds. ASM Press,
Washington DC
Manual of Clinical Microbiology. Fifth edition. 1991. Albert Balows ed., ASM, Washington DC
Prudent Practices in the Laboratory: Handling and disposal of chemicals. 1995 National Research Council. National
Academy Press, Washington
64
Biosafety Manual
Table 5: Summary and Comparison of Liquid Disinfectants (Page 1)
Commonly used disinfectants, recommended when appropriate.
Class
Recommended Use
How They Work
Advantages
70% Isopropyl
alcohol
solution
 Cleaning some
instruments
 Cleaning skin
 Changes protein
structure of
microorganism
 Presence of water
assists with killing action
 Fairly inexpensive
Chlorine
compounds
 Spills of human body
fluids
 Bactericidal - Good
 Fungicidal - Good
 Sporicidal - Good at
>1000 ppm Sodium
Hypochlorite
 Kill hardy viruses
(e.g., hepatitis)
 Kill a wide range of
organisms
 Inexpensive
 Penetrates well
 Relatively quick
microbial kill
 May be used on food
prep surfaces
Gluteraldehyde
 Bactericidal - Good
 Fungicidal - Good
 Tuberculocidal Excellent
 Virucidal - Good
 Sporicidal - Good
 Free available chlorine
combines with contents
within microorganism reaction byproducts cause
its death
 Need 500 to 5000 ppm
 Produce chemical
combination with cell
substances
 Depend upon release of
hypochlorous acid
 Coagulates cellular
proteins
Biosafety Manual
 Non-staining,
relatively non-corrosive
 Useable as a sterilant
on plastics, rubber,
lenses, stainless steel,
and other items that
can’t be autoclaved
Disadvantages
Comments/Hazards
Examples
 <50% solution not very
effective
 Not active when organic
matter present
 Not active against certain
types of viruses
 Evaporates quickly - contact
time not sufficient for killing
 Corrode metals such as
stainless, aluminum
 Organics may reduce activity
 Increase in alkalinity
decreases bactericidal property
 Unpleasant taste and odor
 Tuberculocidal - with
extended contact time
Flammable
Eye Irritant
Toxic
 Follow spill procedure
and dilution instructions
 Make fresh solutions
before use
 Eye, skin, and
respiratory irritant
Corrosive
Toxic
 Bleach
solutions
(sodium
hypochlorite)
 Clorox
 Cryosan
 Purex
 Not stable in solution
 Has to be in alkaline solution
 Inactivated by organic
material
Eye, skin and
respiratory irritant.
Sensitizer
Toxic
 Calgocide
14
 Cidex
 Vespore
65
Table 5: Summary and Comparison of Liquid Disinfectants (Page 2)
Class
Recommended Use
How They Work
Advantages
Iodophors
(iodine with
carrier)
 Disinfecting some
semi-critical medical
equipment
 Bactericidal - Very
Good
 Fungicidal - Excellent
 Virucidal - Excellent
Phenolic
Compounds
 Bactericidal Excellent
 Fungicidal - Excellent
 Tuberculocidal Excellent
 Virucidal - Excellent
Quaternary
ammonium
compounds
(QUATS)
 Ordinary
housekeeping
(e.g., floors,
furniture, walls)
 Bactericidal Excellent
 Fungicidal - Good
 Virucidal - Good (not
as effective as phenols)
 Free iodine enters
microorganism and binds
with its cellular
components
 Carrier helps penetrate
soil/fat
 Need 30 to 50 ppm
 Probably by disorder of
protein synthesis due to
hindrance and/or
blocking of hydrogen
bonding
 Gross protoplasmic
poison
 Disrupts cell walls
 Precipitates cell
proteins
 Low concentrations
inactivate essential
enzyme systems
 Affect proteins and cell
membrane of
microorganism
 Release nitrogen and
phosphorous from cells
Disadvantages
Comments/Hazards
Examples
 Kill broad range of
organisms
 Highly reactive
 Low tissue toxicity
 Kill immediately rather than
by prolonged period of stasis
 Not affected by hard water
 May be used on food prep
surfaces
 May stain plastics or
corrode metal
 May stain skin/laundry
 Stains most materials
 Odor
 Some organic and
inorganic substances
neutralize effect
 Tuberculocidal - with
extended contact time
 Sporicidal - some
 Dilution critical - follow
directions!
 Use only EPAregistered hard surface
iodophor disinfectants
 Don’t confuse skin
antiseptic iodophors for
disinfectants
Skin and eye irritant
Corrosive
Toxic
 Bactergent
 Hy-Sine
 Ioprep
 Providoneiodine;
betadine

Wescodyne
 Nonspecific concerning
bactericidal and fungicidal
action
 When boiling water would
cause rusting, the presence of
phenolic substances produces
an anti-rusting effect
 Unpleasant odor
 Some areas have
disposal restrictions
 Effectiveness reduced
by alkaline pH, natural
soap, or organic
material
 Sporicidal - NO
 Do not eliminate
spores, TB bacteria,
some viruses
 Effectiveness
influenced by hard water
 Layer of soap
interferes with action
Skin and eye irritant
Sensitizer
Corrosive
Toxic
 Hil-Phene
 Lph
 Metar
 Vesphene
 Select from EPA list of
hospital disinfectants
Skin and eye irritant
Toxic
 Coverage
258
 End-Bac
 Hi Tor
 Contain a detergent to help
loosen soil
 Rapid action
 Colorless, odorless
 Non-toxic, less corrosive
 Highly stable
 May be used on food prep
surfaces
Barbara Fox Nellis (8-23-96)
66
Biosafety Manual
Table 6: Summary of Practical Disinfectants
Inactivates
Vegetative Bacteria
Lipoviruses
Nonlipid Viruses
Bacterial Spores
Treatment Requirements
Use Dilution
Contact Time-minutes
Lipovirus
Broad Spectrum
Important Characteristics
Effective Shelf Life
> 1 weekc
Corrosive
Flammable
Explosion Potential
Inactivated by organic
matter
Skin Irritant
Eye Irritant
Respiratory Irritant
Toxicd
Applicability
Waste Liquids
Dirty Glassware
Equipment, Surface
Decontamination
Proprietary Productse
Quatenary Ammonium
Compounds
Phenolic
Compounds
Chlorine
Compounds
Iodophor
Ethyl Alcohol
Isopropyl
Alcohol
Formaldehyde
Glutaraldehyde
+
+
-
+
+a
-
+
+
+
+
+
+
+
+
+
+a
-
+
+
-
+
+
+
+
+
+
+
+
0.1-2.0%
1.0-5.0%
500ppmb
25-1600ppmb
70-85%
70-85%
0.2-0.8%
2%
10
NE
10
NE
10
30
10
30
10
NE
10
NE
10
30
10
30
+
+
-
+
+
+
+
+
+
+
-
+
+
+
+
+
-
+
-
-
-
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
A-33, CDQ, End-Bac,
Hl-Tor, Mikro-Quat
Hil-Phene,
Chloramine
Hy-Sine, Ioprep,
Sterac
Cidex
Metar, MikroT, Clorox,
Mikroklene,
Bac, O-Syl
Purex
Wescodyne
Source: Adapted from Laboratory Safety Monograph, A Supplement to the NIH Guidelines for Recombinant DNA Research. National Institutes of Health, Office of
Research Safety, National Cancer Institute, and the Special Committee of Safety and Health Experts, Bethesda, Maryland. January 1989: 104-105
a
d
+ Yes
Variable results depending on virus
By skin or mouth or both
b
e
- No
Available Halogen
Space limitations preclude listing all products available. Individual listings (or omissions)
c
NE Not Effective
Protected from light and air
do not imply endorsement (or rejection) of any product b y the National Institutes of Health.
Biosafety Manual
67
Table 7: Reprocessing Methods for Equipment Used in the Health Care Setting
Sterilization:
High-Level
Disinfection:
IntermediateLevel
Disinfection:
Low-Level
Disinfection:
Environmental
Disinfection:
Destroys:
All forms of microbial life including high numbers of bacterial spores.
Methods:
Steam under pressure (autoclave), gas (ethylene oxide), dry heat or immersion in an approved chemical “sterilant” (e.g., US Environmental Protection
Agency-approved) for prolonged period of time, e.g., 6-10 hours or according to manufacturer’s instructions. Note: Liquid chemical “sterilants”
should be used only on those instruments that are impossible to sterilize or disinfect with heat.
Use:
For those instruments or devices that penetrate skin or contact normally sterile areas of the body, e.g., scalpels, needles, etc. Disposable invasive
equipment eliminates the need to reprocess these types of items. When indicated, however, arrangements should be made with a health-care facility
for reprocessing of reusable invasive instruments.
Destroys:
All forms of microbial life except high numbers of bacterial spores.
Methods:
Hot water pasteurization (80-100°C, 30 minutes) or exposure to an approved (e.g., US EPA-approved) “sterilant” chemical as above, except for a
short exposure time (10-45 minutes or as directed by the manufacturer).
Use:
For reusable instruments or devices that come into contact with mucous membranes (e.g., laryngoscope blades, endotracheal tubes, etc.).
Destroys:
Mycobacterium tuberculosis, vegetative bacteria, most viruses and most fungi, but does not kill bacterial spores.
Methods:
Approved (e.g. US EPA-approved) “hospital disinfectant” chemical germicides that have a label claim for tuberculocidal activity; commercially
available hard-surface germicides or solutions containing at least 500 ppm free available chlorine (a 1:10 dilution of common household bleachabout ¼ cup bleach per gallon of tap water).
Use:
For those surfaces that come into contact only with intact skin, e.g., stethoscopes, blood pressure cuffs, splints, etc., and have been visibly
contaminated with blood or bloody body fluids. Surfaces must be pre-cleaned of visible material before the germicidal chemical is applied for
disinfection.
Destroys:
Most bacteria, some viruses, some fungi, but no Mycobacterium tuberculosis or bacterial spores.
Methods:
Approved (e.g., US EPA-approved) “hospital disinfectants” (no label claim for tuberculocidal activity).
Use:
Use these excellent cleaning agents for routine housekeeping or removal of soiling in the absence of visible blood contamination.
Environmental surfaces that have become soiled should be cleaned and disinfected using any cleaner or disinfectant that is intended for
environmental use. Such surfaces include floors, woodwork, ambulance seats, countertops, etc.
Important: To assure effectiveness of any sterilization or disinfection process, equipment and instruments must first be thoroughly cleaned of all visible soil.
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Biosafety Manual
Shipment of Biological Materials
The following regulations apply to the packaging and shipment of biological materials:
U.S. Department of Transportation, 49 CFR Parts 171-180 and amendments
U.S. Public Health Service, 42 CFR Part 72, Interstate Shipment of Etiolgic Agents
U.S. Department of Labor, Occupational Safety and Health Administration, 29 CFR Part
1910.1030, Bloodborne Pathogens
International Air Transport Association (IATA), Dangerous Goods Regulations
U.S. Postal Service, 39 CFR Part 111, Mailability of Etiologic Agents, Mailability of Sharps and
Other Medical Devices, and Publication 52, Acceptance of Hazardous, Restricted or
Perishable Matter
International Civil Aviation Organization, Technical Instructions for the Safe Transport of
Dangerous Goods by Air
United Nations, Recommendations of the Committee of Experts on the Transportation of
Dangerous Goods
All North American airlines and FedEx, the largest shipper of infectious materials, use
the IATA regulation (also referred to as the Dangerous Goods Regulation or DGR) as
their standard. Meeting the conditions of this standard will ensure meeting the provisions
of the other US regulations.
General information is provided below, but please contact the EH&S Biological Safety
Office for specific information. Note that for any biological materials for which a state
or federal permit or license is required, registration with the EH&S Biological Safety
Office is also required.
Any NAU faculty/staff, regardless of job or task, must be trained to package, ship or
receive packages of materials/samples classified by regulation as “dangerous goods”.
This training is available from EH&S. Regulation requires retraining every other
year OR whenever the regulations change.
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69
Laboratory Spills
A spill of biological materials that takes place in the open laboratory may create a serious
problem. Every effort should be taken to prevent spilling materials.
A spill poses less of a problem if it happens inside a biological safety cabinet, provided
splattering to the outside of the cabinet does not occur. Direct application of concentrated
liquid disinfectant and a thorough wipe down of the internal surfaces of the cabinet will
usually be effective for decontaminating the work zone, but gaseous sterilants will be
required to disinfect the interior sections of the cabinet and HEPA filter(s).
Spill in the Open Laboratory
Advance preparation for management of a spill is essential. A “spill kit” may include the
following:
 Leak-proof containers
 Forceps
 Paper towels
 Disinfectant
 Respirators, if necessary
 Rubber gloves and other personal protective equipment
 Absorbent powder or other material
 Red bag
If potentially hazardous biological material is spilled in the laboratory, avoid inhaling any
airborne material by holding your breath and leaving the laboratory. Warn others in the
area by posting signs and go directly to wash or change room area. If clothing is known
(or suspected) to be contaminated, remove the clothing with care, folding the
contaminated area inward. Discard the clothing into a bag or place the clothing directly
in an autoclave. Wash all potentially contaminated body areas as well as the arms, face
and hands. Shower if facilities are available. Re-entry into the laboratory should be
delayed for a period of at least thirty minutes to allow reduction of the aerosol generated
by the spill.
Protective clothing should be worn when entering the laboratory to clean the spill area.
Rubber gloves, autoclavable or disposable footwear, an outer garment, and a respirator
equipped with a HEPA filter should be worn. If the spill was on the floor, do not use a
surgical gown that may trail on the floor when bending down. Take the “ spill kit” into
the laboratory room, place a discard container near the spill, and transfer large fragments
of material into it; replace the cover. Using a freshly prepared hypochlorite containing
5000 ppm (10% by volume) available chlorine, iodophor solution containing 1600 ppm
iodine, or other appropriate EPA registered disinfectant, carefully pour the disinfectant
around and into the visible spill. Avoid splashing and work from the outside toward the
center. Allow 30 minutes' contact time. Use paper or cloth towels to wipe up the
disinfectant and spill, working toward the center of the spill. Discard all towels and other
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Biosafety Manual
clean up materials into a discard container as they are used. Wipe the outside of the
discard containers, especially the bottom, with a towel soaked in a disinfectant. Place the
discard container and other materials in an autoclave and sterilize. Alternately, place all
materials in the appropriate biomedical waste system for incarceration. Remove shoes or
shoe covers, outer clothing, respirator, and gloves and sterilize by autoclaving. Wash
hands, arms and face, or if possible, shower.
If gaseous decontamination of the room is required, contact the Biosafety Office.
Spill in a Biological Safety Cabinet
A spill that is confined to the interior of the biological safety cabinet should present little
or no hazard to personnel in the area. However, chemical disinfection procedures should
be initiated at once while the cabinet ventilation system continues to operate to
prevent escape of contaminants from the cabinet.
Spray or wipe walls, work surfaces, and equipment with a disinfectant. A disinfectant
with a detergent has the advantage of detergent activity that will help clean the surfaces
by removing both dirt and microorganisms. A suitable disinfectant is a 3% solution of an
iodophor such as Wescondyne or a 1:100 dilution of household bleach (e.g. Chlorox)
with 0.7% nonionic detergent. The operator should wear gloves during this procedure.
Use sufficient disinfectant solution to ensure that the drain pans and catch basins below
the work surface contain the disinfectant. Lift the front exhaust grill and tray and wipe all
surfaces. Wipe the catch basin and drain the disinfectant into a container. The
disinfectant, gloves, wiping cloth and sponges should be discarded into an autoclave pan
and autoclaved.
The above procedure will not disinfect the filters, blower, air ducts, or other interior parts
of the cabinet. If the entire interior of the cabinet needs to be sterilized, contact the
EH&S Biosafety Office for the name of the current contractor.
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Biological Safety Cabinets
The following is excerpted from Primary Containment for Biohazards: Selection, Installation and Use of
Biological Safety Cabinets , 1995, Centers for Disease Control & Prevention/National Institutes of Health
This chapter presents information on the
selection, function, and use of biological
safety cabinets (BSCs), which are the
primary
means
of
containment
developed for working safely with
infectious microorganisms. BSCs are
designed
to
provide
personnel,
environment, and product protection
when
appropriate
practices
and
procedures are followed.
Three kinds of biological safety
cabinets, designated as Class I, II, and
III, have been developed to meet varying
research and clinical needs.
Biological safety cabinets use high efficiency particulate air (HEPA) filters in their
exhaust and/or supply systems.
Biological Safety Cabinets (BSCs)
The similarities and differences in protection offered by the various classes of biosafety
cabinets are reflected in Table 1.
The Class I BSC
This type of cabinet is not for aseptic or sterile technique. The Class I BSC provides
personnel and environmental protection, but no product protection. It is similar in air
movement to a chemical fume hood, but has a HEPA filter in the exhaust system to
protect the environment.
The Class II BSC
The Class II (Types A and B) biological safety cabinets provide personnel,
environmental, and product protection. Air flow is drawn around the operator into the
front grille of the cabinet, which provides personnel protection. In addition, the
downward laminar flow of HEPA-filtered air provides product protection by minimizing
the chance of cross-contamination along the work surface of the cabinet. Because cabinet
air has passed through the exhaust HEPA filter, it is contaminant-free (environmental
protection), and may be recirculated back into the laboratory (Type A BSC) or ducted out
of the building (Type B BSC).
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The Class II, Type A BSC
An unducted Class II Type A BSC is not to be used for work involving volatile or toxic
chemicals. The buildup of chemical vapors in the cabinet (by recirculated air) and in the
laboratory (from exhaust air) could create health and safety hazards. Generally, BSCs are
not for use with chemicals. Small quantities of chemicals and chemotherapeutic agents
may be used in ducted, 100% BSCs.
It is possible to duct the exhaust from a Type A cabinet out of the building. However, it
must be done in a manner that does not alter the balance of the cabinet exhaust system,
thereby disturbing the internal cabinet air flow. The typical method of ducting a Type A
cabinet is to use a “thimble”, or canopy unit, which maintains a small opening (usually 1
inch) around the cabinet exhaust filter housing. The volume of the exhaust must be
sufficient to maintain the flow of room air into the space between the thimble unit and the
filter housing (contact manufacturers for any additional specifications). The thimble must
be removable or be designed to allow for operational testing of the cabinet. The
performance of a cabinet with this exhaust configuration is unaffected by fluctuations in
the building exhaust system.
“Hard-ducting” (i.e., direct connection) of Class II Type A cabinets to the building
exhaust system is not recommended. The building exhaust system must be precisely
matched to the airflow from the cabinet in both volume and static pressure. However,
fluctuations in air volume and pressure that are common to all building exhaust systems
make it difficult, if not impossible, to match the airflow requirements of the cabinet.
The Class II, Type B1 BSC
Some biomedical research requires the use of small quantities of certain hazardous
chemicals, such as carcinogens. The powdered form of these carcinogens should be
weighed or manipulated in a chemical fume hood or a static-air glove box. Carcinogens
used in cell culture or microbial systems require both biological and chemical
containment.
Type B1 cabinets must be hard-ducted to their own dedicated exhaust system. As
indicated earlier, blowers on laboratory exhaust systems should be located at the terminal
end of the ductwork. A failure in the building exhaust system may not be apparent to the
user, as the supply blowers in the cabinet will continue to operate. A pressureindependent monitor should be installed to sound an alarm and shut off the BSC supply
fan, should failure in exhaust airflow occur. Since all cabinet manufacturers do not
supply this feature, it is prudent to install a sensor in the exhaust system as necessary. To
maintain critical operations, laboratories using Type B BSCs should connect the exhaust
blower to the emergency power supply.
The Class II, Type B2 BSC
This BSC is a total-exhaust cabinet; no air is recirculated within it. This cabinet
provides simultaneous primary biological and chemical containment.
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73
Should the building or cabinet exhaust fail, the cabinet will be pressurized, resulting in a
flow of air from the work area back into the laboratory. Cabinets built since the early
1980's usually have an interlock system installed by the manufacturer to prevent the
supply blower from operating whenever the exhaust flow is insufficient. Presence of
such an interlock system should be verified; systems can be retrofitted if necessary. A
pressure-independent device should monitor exhaust air movement.
The Class II, Type B3 BSC
This biological safety cabinet is a ducted Type A cabinet. All positive pressure
contaminated plenums within the cabinet are surrounded by a negative air pressure
plenum. Thus, leakage in a contaminated plenum will be into the cabinet and not into the
environment.
The Class III BSC
The Class III biological safety cabinet was designed for work with biosafety level 4
microbiological agents, and provides maximum protection to the environment and the
worker. It is a gas-tight enclosure with a non-opening view window.
Long, heavy-duty rubber gloves are attached in a gas-tight manner to ports in the cabinet
and allow for manipulation of the materials isolated inside. Although these gloves restrict
movement, they prevent the user's direct contact with the hazardous materials. The tradeoff is clearly on the side of maximizing personal safety. Depending on the design of the
cabinet, the supply HEPA filter provides particulate-free, albeit somewhat turbulent,
airflow within the work environment.
Horizontal Laminar Flow “Clean Bench”
Horizontal laminar flow clean air benches are NOT BSCs. They discharge HEPA-filtered
air across the work surface and toward the user. These devices only provide product
protection. They can be used for certain clean activities, such as the dust-free assembly of
sterile equipment or electronic devices. These benches should never be used when
handling cell culture materials or drug formulations, or when manipulating potentially
infectious materials. The worker can be exposed to materials (including proteinaceous
antigens) being manipulated on the clean bench, which may cause hypersensitivity.
Horizontal clean air benches should never be used as a substitute for a biological safety
cabinet in research, biomedical or veterinary laboratories and/or applications.
Vertical Laminar Flow “Clean Bench”
Vertical laminar flow clean benches also are NOT BSCs. They may be useful, for
example, in hospital pharmacies when a clean area is needed for preparation of
intravenous drugs. While these units generally have a sash, the air is usually discharged
into the room under the sash, resulting in the same potential problems as the horizontal
laminar flow clean benches.
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Biosafety Manual
Operations within a Class II BSC
Laboratory Hazards
Many common procedures conducted in BSCs may create splatter or aerosols. Good
microbiological techniques should always be used when working in a biological safety
cabinet. For example, techniques to reduce splatter and aerosol generation will minimize
the potential for exposure to personnel from infectious materials manipulated within the
cabinet. Class II cabinets are designed so that horizontally nebulized spores will be
captured by the downward flowing cabinet air within fourteen inches of travel. Therefore,
as a general rule of thumb, keeping clean materials at least one foot away from aerosolgenerating activities will minimize the potential for cross-contamination.
The general workflow should be from “clean to contaminated (dirty).” Materials and
supplies should be placed in such a way as to limit the movement of “dirty” items over
“clean” ones.
Several measures can be taken to reduce the chance for cross-contamination when
working in a BSC. Work at least 6” back from the front edge and never cover the front
grill. Opened tubes or bottles should not be held in a vertical position. Investigators
working with Petri dishes and tissue culture plates should hold the lid above the open
sterile surface to minimize direct impact of downward air. Bottle or tube caps should not
be placed on the toweling. Items should be recapped or covered as soon as possible.
Open flames are not permitted in the near microbe-free environment of a BSC. On an
open bench, flaming the neck of a culture vessel will create an upward air current that
prevents microorganisms from falling into the tube or flask. An open flame in a BSC,
however, creates turbulence that disrupts the pattern of air supplied to the work surface.
When deemed absolutely necessary, touch-plate microburners equipped with a pilot light
to provide a flame on demand may be used. Internal cabinet air disturbance and heat
buildup will be minimized. The burner must be turned off when work is completed.
Small electric “furnaces” are available for decontaminating bacteriological loops and
needles and are preferable to an open flame inside the BSC. Disposable sterile loops
should be used to eliminate the need for heat or flame.
Aspirator bottles or suction flasks should be connected to an overflow collection flask
containing appropriate disinfectant, and to an in-line HEPA or equivalent filter. This
combination will provide protection to the central building vacuum system or vacuum
pump, as well as to the personnel who service this equipment. Inactivation of aspirated
materials can be accomplished by placing sufficient chemical decontamination solution
into the flask to kill the microorganisms as they are collected. Once inactivation occurs,
liquid materials can be disposed of appropriately as noninfectious waste.
Investigators must determine the appropriate method of decontaminating materials that
will be removed from the BSC at the conclusion of the work. When chemical means are
appropriate, suitable liquid disinfectant should be placed into the discard pan before work
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75
begins. Items should be introduced into the pan with minimum splatter, and allowed
appropriate contact time as per manufacturer's instructions. Alternatively, liquids can be
autoclaved prior to disposal. Contaminated items should be placed into a biohazard bag
or discard tray inside the BSC. Water should be added to the bag or tray prior to
autoclaving.
When a steam autoclave is to be used, contaminated materials should be placed into a
biohazard bag or discard pan containing enough water to ensure steam generation during
the autoclave cycle. The bag should be taped shut or the discard pan should be covered in
the BSC prior to removal to the autoclave. The bag should be transported and autoclaved
in a leak-proof tray or pan.
Decontamination
Surface Decontamination
All containers and equipment should be surface decontaminated and removed from the
cabinet when work is completed. At the end of the workday, the final surface
decontamination of the cabinet should include a wipe-down of the work surface, the
cabinet's sides and back, and the interior of the glass. If necessary, the cabinet should
also be monitored for radioactivity and decontaminated when necessary. Investigators
should remove their gloves and gowns and wash their hands as the final step in safe
microbiological practices.
Small spills within the BSC can be handled immediately by removing the contaminated
absorbent paper toweling and placing it into the biohazard bag. Any splatter onto items
within the cabinet, as well as the cabinet interior, should be immediately wiped with a
towel dampened with decontaminating solution. Gloves should be changed after the work
surface is decontaminated and before placing clean absorbent toweling in the cabinet.
Hands should be washed whenever gloves are changed or removed.
Spills large enough to result in liquids flowing through the front or rear grilles require
more extensive decontamination. All items within the cabinet should be surface
decontaminated and removed. After ensuring that the drain valve is closed,
decontaminating solution can be poured onto the work surface and through the grille(s)
into the drain pan.
Thirty minutes is generally considered an appropriate contact time for decontamination,
but this varies with the disinfectant and the microbiological agent. Manufacturer's
directions should be followed. The spilled fluid and disinfectant solution on the work
surface should be absorbed with paper towels and discarded into a biohazard bag. The
drain pan should be emptied into a collection vessel containing disinfectant. A flexible
tube should be attached to the drain valve and be of sufficient length to allow the open
end to be submerged in the disinfectant within the collection vessel. This procedure
serves to minimize aerosol generation. The drain pan should be flushed with water and
the drain tube removed.
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Biosafety Manual
Should the spilled liquid contain radioactive material, a similar procedure can be
followed. The EH&S Radiation Safety Officer should be contacted for specific
instructions. See contact information in the front of this manual.
Gas Decontamination
BSCs that have been used for work involving infectious materials must be
decontaminated before HEPA filters are changed, internal repair work is done or before a
BSC is relocated. The most common decontamination method uses formaldehyde gas,
although more recently, hydrogen peroxide vapor has been used successfully. This
environmentally benign vapor is useful in decontaminating HEPA filters, isolation
chambers, and centrifuge enclosures. Call the EH&S Biosafety Office for the current BSC
vendor who does the gas decontaminations and certifications. All BSCs must be recertified following any gas decontamination, maintenance or relocation.
Engineering Requirements
Ultraviolet Lamps
Ultraviolet (UV) lamps are not required in BSCs. If installed, UV lamps must be cleaned
weekly to remove any dust and dirt that may block the germicidal effectiveness of the
ultraviolet light. The lamps should be checked periodically with a meter to ensure that
the appropriate intensity of UV light is being emitted. UV lamps must be turned off when
the room is occupied to protect eyes and skin from UV exposure, which can burn the
cornea and cause skin cancer. Do not depend on UV lamps to disinfect the area.
BSC Placement
The ideal location for the biological safety cabinet is remote from the entry (e.g., the rear
of the laboratory away from traffic), since people walking parallel to the face of a BSC
can disrupt the air curtain. The air curtain created at the front of the cabinet is quite
fragile, amounting to a nominal inward and downward velocity of 1 mph. Open
windows, air supply registers, or laboratory equipment that creates air movement (e.g.,
centrifuges, and vacuum pumps) should not be located near the BSC. Similarly, chemical
fume hoods must not be located close to BSCs.
HEPA Filters
HEPA filters, whether part of a building exhaust system or part of a cabinet, will require
replacement when they become so loaded that sufficient air flow can no longer be
maintained. Filters must be decontaminated before removal.
Certification of BSCs
All BSCs must be certified (according to a National Sanitation Foundation standard)
annually according to NAU policy. Please contact the EH&S Biological Safety Office for
the name and phone number of the current contractor performing this service. Prices and
quality vary widely, so only BSO-approved contractors may be used.
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77
3 – Programs
Animal Contact Medical Monitoring Program
The NAU Animal Contact Medical Monitoring Program is designed to protect employees
from occupational exposure to conditions that may result in animal related illnesses.
According to the Public Health Service, and occupational health program is required for
institutions that employ personnel who have animal contact. The NAU Animal Contact
Monitoring Program is overseen by the NAU Institutional Animal Care and Use
Committee (IACUC).
Occupational Health and Safety (Animal Contact Medical Monitoring Program)
Occupational health and safety is of utmost importance to the NAU IACUC. Regulations
require that the IACUC monitor personnel occupational health programs and training
related to animal use at NAU. Occupational health programs must include training in
basic safety techniques and information about hazards which will be encountered in each
particular job. Hazardous materials must be stored safely and in compliance with MSDS
and OSHA regulations. Posted signs must warn of hazards present. The NAU program is
designed to protect all personnel from occupational exposure to conditions that may result
in animal related illnesses.
Responsibilities Under the Program
All individuals who engage in animal-related work will participate in the university
occupational health program. As the initial step in this program, individuals will
complete the "Occupational Health Questionnaire" found at the IACUC website
www.nau.edu/iacuc. The University Veterinarian will review the questionnaire, make
any comments which may highlight animal work related concerns, and forward the form
to health care providers at Fronske Health Center. At the discression of Fronske medical
personnel, the individual will be scheduled for a brief interview or examination, where
potential health-risks will be discussed relative to the individual's health status and job
requirements. All individuals who work with animals will be current on their tetanus
vaccine. Serology and/or vaccination for rabies, Hepatitis B, toxoplasmosis, measles, and
skin testing for tuberculosis may be required for work with certain species or protocols.
Vaccination for rabies will be required to work with wild or random-source bats, skunks,
and carnivores. Tuberculosis testing will be required for work with all non-human
primates. This program is provided at no cost to the employee.
Principle Investigators are responsible for ensuring that all personnel (including
employees, students, colleagues, collaborators, and volunteers) involved with their
IACUC-approved project are given this program information and that they receive the
appropriate immunizations and tests, etc. Investigators who do not respond to requests
for registration may have their animal use approval rescinded by the IACUC.
Supervisors are responsible for providing information to animal care technicians,
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Biosafety Manual
veterinarians, and associated animal workers (e.g. workers in Animal Care
Services) who work with animals but are not on a specific research project.
The University Veterinarian and/or health care providers at Fronske will conduct an
educational component of the occupational health program as appropriate. Topics to be
discussed include zoonoses relative to the protocol, allergy risk, possible risks to
immunocompromised individuals, and use of protective equipment and supplies such as
gloves, eye protection, masks, lab coats, scrubs, respirators, head covers, boots, and heat
resistant gloves chemical resistant gloves.
An animal exposure case involves mucous membrane contact with animal fluids or
excreta, ingestion of animal fluids or excreta, bites, scratches, etc.. Emergency medical
care for minor animal exposure cases is available at Fronske Health Center during
business hours, Monday-Friday. For more serious cases and during other hours, care is
available that the Flagstaff Medical Center Emergency Room. In cases where individuals
may be exposed to non-routine or exotic diseases, it is necessary to have emergency care
sheets to provide basic information for medical personnel. Such information may include
MSDS's, sheets copied from the "Control of Communicable Diseases Manual," or
monographs written by experts in diseases such as Cercopithicine Herpes Virus. Such
sheets should be readily available so affected individuals may easily take them when
seeking emergency care. MSDS's for chemicals used in laboratories must also be readily
available.
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NAU Bloodborne Pathogen Program
In December 1991, OSHA published the final rule covering
occupational exposure to bloodborne pathogens. This was
adopted by the state of Arizona and written into the Arizona
Administrative Code in January 1993. NAU instituted its
program in the spring of 1993.
The rule requires that those who handle human blood or other potentially infectious
(human) materials as part of their job duties participate in an employer-generated
program. This program shall include development and annual review of a site specific
Exposure Control Plan, annual training regarding exposures, offer of hepatitis B
vaccinations free-of-charge, and post-exposure health care services.
Northern Arizona University manages this program through the EH&S Biological Safety
Office. The BBP program materials and information is available on our website at
www4.nau.edu/regulatorycompliance
In addition to the program materials, the EH&S Biosafety Office provides training
sessions annually.
Hepatitis B vaccinations are given by the Fronske Health Center. Official program
medical records are kept by Fronske Health Center. There is a requirement that Flagstaff
Medical Center staff or other providers send vaccination and post-exposure records to
Fronske Health Center for record-keeping purposes. Declination statements from those
that decline the vaccination series are kept by The EH&S Biosafety Office.
The EH&S Biosafety Office monitors for NAU compliance by requiring training and
vaccination documentation and by confirming BBP participation during the annual
laboratory safety survey which is conducted in each laboratory each year.
For further information, please call the EH&S Biological Safety Office.
BBP Exposure Information
For exposure incidents more than 1 hour away from Flagstaff, go to the nearest medical
facility.
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4 -Medical Surveillance
HIV Research Laboratory
Occupational Medicine Policy
This policy is designed to protect employees who conduct research
with HIV. Researchers who handle, manipulate, or assay live HIV
cultures are covered under this policy.
Pre-employment
Prior to beginning work in an HIV research lab, an employee shall
be given, at no cost to the employee, the following:
1.
2.
3.
A baseline serum sample shall be stored with the NAU Student Health Care Center. This is a
requirement to work in the HIV lab.
A confidential HIV test shall be offered to the employee. The HIV test is available at the NAU
Student Health Care Center. The test must be offered by the employer, but may be declined by
the employee. This test shall include HIV counseling. Results of the test shall be given to the
employee in person in a face-to-face meeting with a health care professional. No exceptions.
All results of HIV testing shall remain completely confidential and shall be stored in a separate
section in the employee’s medical record. At no time, will the employer or any administrator or
official of NAU have access to the employee’s confidential record concerning HIV testing.
If the HIV researcher also works with human blood or other potentially infected material, all
aspects of the NAU Bloodborne Pathogen Program under the OSHA Bloodborne Pathogen
standard shall be implemented for that employee. The employer shall offer free HBV
immunization, annual training, and have a written exposure control plan in the workplace.
Continuing employment
HIV researchers shall be given the following at no cost to the employee:
1.
Annual serum banking shall be offered to the employee, but may be declined.
2.
Annual HIV tests shall be offered to the employee, but may be declined.
3.
Annual training regarding HIV and post-exposure prophylaxis, including the risks of
chemoprophylaxis, shall be given to the employee. This is a required, not optional, component.
Post-exposure prophylaxis
An exposure to an HIV culture in a research laboratory is considered a “highest risk exposure”
according to Public Health Service CDC guidelines. The following shall be implemented:
1.
If the exposure is to intact or broken skin, or in the event of a puncture wound, immediately
wash the affected area with water for five minutes. Soap may be used, if immediately available.
If the exposure is to the eyes, they shall be rinsed for five minutes in an eyewash station. Other
exposed mucous membranes (nose and mouth) shall be rinsed for five minutes with water.
2.
The employee shall proceed immediately to the nearest health care facility. It is important for
the employee to seek treatment within the first two hours of exposure.
3.
The employee may be seen at the Fronske Health Center between the hours of 8:00 a.m. and
4:00 p.m. Monday through Friday. After hours, employees should proceed to the Flagstaff
Medical Center Emergency Department, where they will be seen immediately, free-of-charge.
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4.
5.
6.
7.
8.
9.
The current Public Health Service (CDC) chemoprophylactic treatment/ post-exposure
prophylaxis (PEP) shall be recommended and offered to an employee with the highest risk
exposure, within the first two hours of exposure. However, medical use of chemoprophylaxis
is a decision that shall be made by a health care professional in conjunction with the employee,
based upon the most current CDC recommendations, the nature of the exposure event, and
other medical factors. Flagstaff Medical Center pharmacy is well stocked with the appropriate
medication.
Baseline tests and informed consent statements shall be required for PEP.
Counseling concerning the risks and benefits of the chemoprophylactic treatment shall be given
to the employee at the time of the exposure event.
The employee may contact an infectious disease specialist, if desired.
All HIV exposures that occur in the workplace are covered under Worker’s Compensation.
Flagstaff Medical Center and NAU personnel are familiar with the paperwork to process this
type of Worker’s Compensation claim for NAU employees.
Post-exposure follow-up, including the offer of HIV testing and counseling, shall be given in
accordance with the CDC recommendations and the OSHA Bloodborne Pathogen rule. The
Flagstaff Medical Center protocol on HIV exposures will be followed.
References
NAU Bloodborne Pathogen Program Compliance Materials; www4.nau.edu/regulatory compliance
OSHA. 29 CFR Part 1910.1030. Occupational Exposure to Bloodborne Pathogens, Final Rule, Dec.
6, 1991
CDC. Update: Provisional public health service recommendations for chemoprophylaxis after
occupational exposure to HIV; MMWR 1996; 45(22):468-472
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Immunoprophylaxis
The Northern Arizona University follows the recommendations of the Centers for Disease Control and
Prevention (CDC) and the Public Health Service Advisory Committee for Immunization Practices
(ACIP) for vaccination of at-risk personnel.
Currently, NAU has written policies regarding the need for hepatitis B, anthrax, plague, tetanus, rabies,
and vaccinia vaccination for certain at-risk personnel. Other vaccinations may be required or
recommended, as needed. Particular attention is given to individuals who are or may become
immunocompromised, as recommendations for vaccine administration may be different than for
immunologically competent adults. Various other factors may be taken into account such as
pregnancy, history of allergy, or HIV status.
When considering the need for immunization, a risk assessment will be conducted by the health care
provider in conjunction with information regarding the experimental agent provided by the EH&S
Biological Safety Office.
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Recommendations for Prophylactic Immunization of Laboratory Personnel
Working with Infectious Agents
Please note, the following is derived from the CDC publication, Biosafety in Microbiological and
Biomedical Laboratories, 5th Edition, 2007 and the CDC publication, “Update on Adult Immunization:
Recommendations of the Immunization Advisory Committee (ACIP),” 1991, MMWR vol. 40.
Bacterial agents
1. Bacillus anthracis
A licensed vaccine is available through the CDC. However, immunization of laboratory
personnel is not recommended unless frequent work with clinical specimens or diagnostic cultures
is anticipated and in settings in which production quantities, high concentrations, or procedures
with a high potential for aerosol (all BSL-3 work) are encountered. In these settings, vaccination
is recommended for all persons working with the agent, all persons working in the laboratory
room where the cultures are handled, and all persons working with infected animals.
2. Clostridium botulinum
A pentavalent (ABCDE) botulism toxoid is available through the CDC as an investigational new
drug (IND). This toxoid is recommended for personnel working with cultures of C. botulinum or
its toxins.
3. Clostridium tetani
The adult diphtheria-tetanus toxoid is highly recommended for laboratory personnel who
manipulate cultures or toxins and is required by the NAU Animal Contact Program for all NAU
personnel who work with animals. This immunization should be given every 10 years.
4. Corynebacterium diphtheria
The adult diphtheria-tetanus toxoid every 10 years is recommended for laboratory personnel who
work with this agent.
5. Francisella tularensis
Vaccination for F. tularensis is available and should be considered for personnel working with
infectious material or infected rodents. It is recommended for for persons working in or entering
laboratories or animal rooms where cultures or animals are maintained.
6. Neisseria meningitidis
Vaccines for N. meningitidis are available and should be considered for personnel regularly
working with infectious materials. The reader is advised to consult the current recommendations
of the Advisory Committee on Immunization Practices (ACIP) published in the CDC Morbidity
and Mortality Weekly Report (MMWR) for recommendations for vaccination against N.
meningitidis.
7. Salmonella typhi
Vaccines for S. typhi are available and should be considered for personnel regularly working with
potentially infectious materials. The reader is advised to consult the current recommendations of
the Advisory Committee on Immunization Practices (ACIP) published in the CDC Morbidity and
Mortality Weekly Report (MMWR) for recommendations for vaccination against S. typhi.
8. Yersinia pestis
Vaccination for Y. pestis is available and should be considered for personnel working with
infectious materials or infected rodents. For example: laboratory personnel who routinely perform
procedures that involve viable Y. pestis; and persons (e.g., mammalogists, ecologists, and other
field workers) who have regular contact with wild rodents or their fleas in areas in which plague is
enzootic or epizootic. The reader is advised to consult the current recommendations of the
Advisory Committee on Immunization Practices (ACIP) published in the CDC Morbidity and
Mortality Weekly Report (MMWR) for information on vaccination against Y. pestis.
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Rickettsial agents
1.
Coxiella burnetii
An investigational Phase I, Q fever vaccine (IND) is available on a limited basis from the Special
Immunizations Program (301-619-4653) of the USAMRIID, Fort Detrick, Maryland, for at-risk
personnel under a cooperative agreement with the individual's requesting institution. The use of
this vaccine should be restricted to those who are at high risk of exposure and who have no
demonstrated sensitivity to Q fever antigen.
Viral agents
1.
Hepatitis A virus
A vaccine is available and recommended for those who work with hepatitis A virus.
2.
Hepatitis B virus
A vaccine is available and highly recommended for those who work with hepatitis B virus. The
hepatitis B vaccine is recommended for those who have occupational exposure to human blood
or other potentially infectious human materials, according to the OSHA rule and the NAU
Bloodborne Pathogen Program.
3.
Poliovirus
All laboratory personnel who work directly with the agent are required to have documented
polio vaccination or demonstrated serologic evidence of immunity to all three poliovirus types.
4.
Poxviruses: vaccinia, cow pox, monkey pox
Vaccination is required for those who work with or around orthopox viruses.
5.
Rabies virus
Immunization is recommended for all individuals prior to work with rabies virus or infected
animals, or engaging in diagnostic, production, or research activities with rabies virus.
Immunization is also recommended for all individuals entering or working in the same room
where rabies virus or infected animals are used. The NAU Animal Contact Program requires
immunization for all employees (e.g. veterinarians and animal control personnel) who come
into contact with unvaccinated carnivores.
6.
Arboviruses
For the following arthropod-borne viruses there is a vaccine available and it is recommended
for those who work with the virus.
Eastern equine encephalitis
Junin
Western equine encephalitis
Mucambo
Cabassue
Rift Valley Fever
Chikungunya
Venezualan equine encephalitis
Everglades
Yellow Fever
7.
Measles, mumps, and German measles virus (Rubeola, paramyxovirus, and rubella virus)
Immunizations are usually given in childhood. Vaccines are highly recommended for all
unvaccinated or serologically negative adults.
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Vaccinia Immunization Policy
All personnel who work in an area where vaccinia, orthopoxviruses, or animals vaccinated with those
agents are used, handled or housed must have received a vaccinia vaccination within the last ten years.
Individuals must declare their vaccination status prior to engaging in such work. This may be
accomplished by using the NAU Smallpox Vaccine Consent form.
To fulfill this requirement:
1.
Request a Vaccinia Vaccination Information Packet from the EH&S Biosafety Office.
2.
Following review of the material, questions should be directed as follows:
a.
b.
Medical questions – Fronske Health Center
Virology questions – BSO and/or principal investigator.
3.
All projects involving “pox” viruses require vaccination.
4.
Any medical condition precluding vaccination will also preclude participation in a project
involving infectious orthopoxvirus.
It shall be the responsibility of each principal investigator to insure that:
1.
Each project is evaluated to determine whether vaccination is required. These evaluations are
conducted by the EH&S Biosafety Office and its various committees.
2.
Each participant complies with the vaccine policy requirements prior to participation in any
covered research project.
3.
The EH&S Biosafety Office is notified of vaccination dates for all researchers working with
vacccinia.
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Health Surveillance for Personnel Working with Infectious Agents
A health surveillance program is available for laboratory personnel who use agents that require it.
Laboratory personnel should receive immunizations (see chapter entitled “Immunoprophylaxis”), such as
hepatitis B vaccination, and medical tests, such as tuberculosis skin testing, when appropriate. The
Animal Contact and Bloodborne Pathogen Programs (see appropriate chapters) provide for health
assessments, risk assessments, medical tests, and immunizations for certain at-risk personnel.
Blood Serum Sampling
For those laboratory personnel working with certain agents under BSL-2 or BSL-3, baseline serum
samples may be appropriate (see chapter entitled “Biological Safety Levels”). The collection and
maintenance of baseline serum samples provide a tool for monitoring serological changes that may
result from the employee’s work. For example, staff who work with non-human primates may be
required to bank a blood sample.
The EH&S Biological Safety Office, with advice from the appropriate health care professional, will
provide information regarding the need for, and the frequency of, blood serum sampling on a case-bycase basis. The need for serum sampling will be dependent upon the agent used and the type of
research in which an individual is engaged. For example, researchers who work with large quantities
of HIV are required to have serum sampling monitored annually and upon accidental exposure.
Health Assessments
The NAU Occupational Medicine Program provides for pre-placement and other health assessments depending
upon the type of work in which an employee will be engaged.
Laboratory personnel who work with agents that are transmitted by aerosol or with certain chemicals or
acute toxins may require a respirator at some stage of the research project. The EH&S Biosafety Office
Occupational Medicine Respiratory Protection Program provides for health assessments that will
determine an employee’s fitness for respirator use.
The EH&S Biosafety Office and Occupational Medicine Program also includes the Hearing
Conservation Program, the Asbestos Monitoring Program, as well as the Animal Contact Program,
discussed in a separate section. Occasionally, researchers who work with infectious agents will require
a health assessment based on the requirements of one of these other programs.
Exposure to Mycobacterium Tuberculosis
All personnel who can be occupationally exposed to either TB research or patients with active TB
should have a yearly TB test for health surveillance.
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5 – Appendices - Information for Researchers
Bio-Agent / R-DNA –
NIH guidelines
Transgenic Animals
ACUTE TOXIN
PROJECT ADDENDUM
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