There are four biosafety levels that provide increasing levels of containment for biological agents. Biosafety level 1 is for work with well-characterized agents that do not cause disease in healthy adults. It requires standard microbiological practices and personal protective equipment like lab coats and gloves. Biosafety level 2 involves agents that pose moderate hazards and requires all BSL-1 practices plus additional containment like restricted access and use of biosafety cabinets. Biosafety level 3 is for serious and potentially lethal agents and requires BSL-2 practices plus additional containment like controlled access, required personal protective equipment, and specialized facility design with airflow controls and separate rooms. Biosafety level 4 is the highest level and involves dangerous and exotic agents

1. M S S A A J I D A S U LTA A N A M A H U S O O K
BIOSAFETY

2. Biosafety
Biosafety is the application of safety precautions that
reduce a laboratorian’s risk of exposure to a potentially
infectious microbe and limit contamination of the work
environment and, ultimately, the community.

3. Biosafety Levels (BSLs)
There are four biosafety levels. Each level has specific controls for
containment of microbes and biological agents.
• The primary risks that determine levels of containment are
infectivity, severity of disease, transmissibility, and the nature of
the work conducted. Origin of the microbe, or the agent in
question, and the route of exposure are also important.
• Each biosafety level has its own specific containment controls
that are required for the following:
Laboratory practices
Safety equipment
Facility construction

4. Biosafety Level 1
• Laboratories suitable for work involving well-
characterized agents that do not cause disease in
healthy humans and are of minimal potential hazard to
laboratory personnel and the environment.
• Examples include:
Bacillus subtilis
Bacillus cerculans
Escherichia coli (non-pathogenic strains of E.
coli such as K-12)
Murine cell lines
Samples from lab mammals

5. Specific considerations for a BSL-1 laboratory include the following:
Laboratory practices
• When working at BSL-1, workers must follow all standard
microbiological practices, receive all specific training on procedures and
equipment.
• Work can be performed on an open lab bench or table.
• Laboratories must be locked when unoccupied. All agents must be
secured against accidental exposure, unauthorized use, and theft. All
recombinant nucleic acids must be stored in locked containers.
Safety equipment
• Personal protective equipment , (lab coats, gloves, eye protection) are
worn as needed.
Facility construction
• A sink must be available for hand washing.
• The lab should have doors to separate the working space with the rest
of the facility.

6. Biosafety Level 2
BSL-2 include the microbes that pose moderate hazards to
laboratorians and the environment. The microbes are
typically indigenous and associated with diseases of
varying severity.
Examples include:
• Staphylococcus aureus,
• Human samples and cell lines,
• Aspergillus fumigatus,
• Toxoplasma gondii,
• Salmonella typhimurium and
• Influenza A.

7. Laboratory practices
• Laboratory personnel should have specific training in
handling pathogenic agents and are supervised by
scientists competent in handling infectious agents and
associated procedures. The Approval Holder must
ensure that laboratory personnel demonstrate
proficiency in standard and special microbiological
practices before working with BSL-2 agents.
• All procedures in which infectious aerosols or splashes
may be created are conducted in biological safety
cabinet (BSCs) or other physical containment
equipment.
• Access to the laboratory is restricted when work is being
conducted.

8. Safety equipment
• At BSL-2, workers are required to follow the standard
microbiological practices and wear the minimum personal
protective equipment (PPE) of a lab coat and gloves.
Additional PPE may be required.
• An autoclave or an alternative method of decontamination is
available for proper disposals.
Facility construction
• The laboratory has self-closing doors.
• A sink and eyewash are readily available.
• All agents must be secured against accidental exposure,
unauthorized use, and theft. All recombinant nucleic acids and
BSL-2 agents must be stored in locked containers. All material
in the open bay or common use areas must be secured when
not in use.

9. Biosafety Level 3
Biosafety Level 3 work involves agents that may cause
serious and potentially lethal infection. The microbes
present can be either indigenous or exotic, and can cause
serious or potentially lethal disease through respiratory
transmission.
• Examples : Mycobacterium tuberculosis, the bacteria
that causes tuberculosis, Coccidioides immitis and
posadasii, Mycobacterium tuberculosis, Chikungunya
Virus and West Nile Virus.
• The primary routes of exposure to personnel working
with these agents are: inhalation, auto inoculation, and
ingestion. Respiratory transmission is the inhalation
route of exposure.

10. Containment requirements:
Laboratory practices
• Laboratorians are under medical surveillance and might receive
immunizations for microbes they work with.
• Access to the laboratory is restricted and controlled at all times.
Safety equipment
• Appropriate PPE must be worn, and respirators might be required.
• All work with microbes must be performed within an appropriate BSC.
Facility construction
• A hands-free sink and eyewash are available near the exit.
• Exhaust air cannot be recirculated, and the laboratory must have
sustained directional airflow by drawing air into the laboratory from
clean areas towards potentially contaminated areas.
• Entrance to the lab is through two sets of self-closing and locking doors.

11. • BSL-3 workers must be fit tested for an appropriate respirator
and individuals wearing contact lenses must wear eye
protection.
• The Approval Holder must monitor and authorize access of all
individuals entering the BSL-3 laboratory. Visitors and
maintenance personnel who enter the BSL-3 laboratory must
be fully informed of the potential risks, required practices and
procedures that they must follow. They must be instructed
about the signs and symptoms of any and all biohazardous
materials manipulated or stored in the laboratory and sign a
statement that they understand the risks.
• Laboratories must be locked when unoccupied. All agents
must be secured against accidental exposure, unauthorized
use, and theft. All recombinant nucleic acids and BSL-3
agents must be stored in locked containers.

12. Biosafety Level 4
• BSL-4 is the highest level of biological safety. There are
a small number of BSL-4 labs in the United States and
around the world. The microbes in a BSL-4 lab are
dangerous and exotic, posing a high risk of aerosol-
transmitted infections. Infections caused by these
microbes are frequently fatal and without treatment or
vaccines.
Examples include
• Ebola
• Marburg viruses etc.

13. • In addition to BSL-3 considerations, BSL-4 laboratories have
the following containment requirements:
Laboratory practices
• Change clothing before entering.
• Shower upon exiting.
• Decontaminate all materials before exiting.
Safety equipment
• All work with the microbe must be performed within an
appropriate Class III BSC , or by wearing a full body, air-
supplied, positive pressure suit.
Facility construction
• The laboratory is in a separate building or in an isolated and
restricted zone of the building.
• The laboratory has dedicated supply and exhaust air, as well
as vacuum lines and decontamination systems.

15. Reference:
• https://rgw.arizona.edu/compliance/rlss/biosafety-
program/biosafety-containment-levels
• https://consteril.com/biosafety-levels-difference/
• https://www.ncbi.nlm.nih.gov/books/NBK218631/
• https://www.cdc.gov/training/QuickLearns/biosafety/
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