BIOSAFETY LEVELS FOR MICROORGANISMS

1. Biosafety levels for
Microorganisms
SUBMITTED BY:-
Ritu sheoran
MSC(Microbiology)
Roll-no-190000304014

2. Biosafety :- safety from potentially infectious materials
of environment and community.
Biosafety cabinets:-
 A biological safety cabinet is a ,ventilated enclosure offering
protection to the user ,the product and the environment from
aerosols arising from the handling of potentially hazardous
microorganisms.
Why we need:-
 Processing of infectious samples to reduce risk of spreading infection.
History :-
 Separate room- reduce contamination
 Hood or steal box – reduce contamination
 Ventilated cabinet- reduce exposure
 HEPA filter – reduce exposure to environment
HEPA filter- High Efficiency Particulate Air Filter
The continuous air flow is discharged to the atmosphere via a HEPA filter.

3. Purpose of Biosafety cabinet:-
 The three States of protection:-
 Personal protection from harmful agents within the cabinet .
 Product protection to avoid contamination of the samples.
 Environment protection from contaminants contained within the cabinet.
Classes of biosafety cabinets:-
Class -1 biosafety cabinet
Class-II biosafety cabinet
Class- III biosafety cabinets

4. Class -1 Biosafety cabinet:-
 Provide good operator protection and environmental
protection but not protect material/ product
protection.
 The inward flow of air can contribute to
contamination of samples.
 Inward airflow is maintained at a minimum velocity of
0.38 m/s .
 Class -1 BSC is designed for general microbiological
research with low and moderate risk agents.
 Biosafety level- 1,2,3.

5. Class-2 biosafety cabinets:-
 Provide personal protection , product and environmental
protection.
 It is commonly found in clinical and research laboratories
working with infectious agents in risk group 2,3 and 4.
 There are four types :- A1,A2,B1,B2.
 The main difference between the types are the ratio of air
exhausted from the BSC to the air that is recirculated within
the BSC ,and the type of exhaust system present.
Reduce cross contamination
A1- 30%out and 70 recirculate
A2-Hepa filter testing /no leakge
B1- 70 out 30 recirculate
B2- total out 100%

6. Class- II Type A1:-
 It has positive pressured plenum.
 No toxic chemical is handled in this type .
 Reduced cross contamination:- 30%of air – exhaust
HEPA filter
 70%of air – recirculates through HEPA filter back
into the work zone of the cabinet.
 Suitabt for working with the agents with biosafety
level 1,2,3 agents.
 Filtration system has two HEPA filters ,one located
on work surface ,the second on the extraction
system which may or may not be connected to
exterior.
 If they connected to exterior,it utilizes a bell type
connection.

7. Type A2:-
 Provide protection ,operator,product ,
environment
 Air velocity entering the cabinet is 50.8 cm/s.
 It has both positive and negative pressured
plenum ,in case of air leaking out of positive
pressured plenum,it is trapped by negative
pressured plenum.
 Suitable for working with agents with biosafety
level 1,2,3.
 Filtration system has 2 HEPA filter .

8. B1 type:-
 For hazardous chemicals and carcinogens.
 Suitable for working with agents with biosafety level 1,2,3.
 70% air out and 30 % recirculate

9. Type B2:-
 Toxic chemicals can be handled in this type .
 100% air is exhausted.
 No air recirculation.
 Product of biosafety level 2,3are used in this type.

10. Class – III Types(“ glove box”):-
 Class III biological safety cabinet is most suitable
for work with biohazardous agents requiring high
contain / biosafety level 3 or 4 .
 Mainly designed for working with biosafety level 4
highly infectious agents and provides the highest
level of protection for the environment , product
and user.
 Completee enclosed ,HEPA filter ventilation
cabinet fitted with glove port and
decontamination capabilities for entry and exit of
material.
 Intake and extraction elements are conducted
through a double door pass through the box.
 Supply and exhaust air – HEPA negative pressure
cabinet ,long heavy duty rubber gloves attached in
a gas right manner to Port in the cabinets.
 2 HEPA filter before exhaust / incineration.

11. Biosafety levels:-
Biosafety :-is the prevention of large scale loss of biological integrity ,
focusing both on ecology and human health.
 A Biosafety level is a set of biocontainment precautions required to isolate
dangerous biological agents in an enclosed laboratory facility.
 The levels of containment range from the lowest biosafety level 1 (BSL-1)to
the highest at level (BSL-4).
 The centres for Disease Control and Prevention (CDC) sets BSL lab levels as a
way of exhibiting specific controls for the containment of microbes and
biological agents.
 Each BSL lab level builds upon on the previous level – thereby creating layer
upon layer of constraints and barriers.

12. CDC biosafety levels:-

14. BSL 1 ( Basic teaching, Research):-
 Biosafety 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.
 Example of a microbe that is typically worked with at a BSL -1 is a non-
pathogenic strain of E.coli, Bacillus subtilis.
 Work is generally conducted on open bench tops using standard
microbiological practices and without the use of special containment
equipment or facility design not required,class -1 may be used.
 Only standard practices required at this level:-
. Frequent hand washing ,after
removing gloves and before leaving lab.
 Door that can be kept closed when working .
 No smoking , eating , drinking, storage of food in laboratory.
 Mechanical pipetting only ( no mouth pipetting allowed).
 “sharp” precautions , including special containers for disposing of needles and
other sharp objects.
 Daily decontamination of all work surfaces when work is complete.

15. Personal protective equipment :-
 Eye protection person who wear contact lenses in laboratories
should also wear goggles or a face shield.
 Gloves and a lab coat or gown .
Laboratoray facilities :- each laboratory contains a sink for
handwashing .
 Laboratory is designed so that it can be easily cleaned.
 Laboratory furniture is sturdy .spaces between benches , cabinets
and equipment are accessible for Cleaning.
 Work surface decontaminated at least once a day and after any spill
of viable material.
 All culture ,stocks and other regulated wastes are decontaminated
before disposal by an approved decontamination method.

16. BSL -2 ( Primary health services, diagnostic
service,research):-
 Agents include :- Salmonella species , Clostridium
botulinum, Hepatitis B virus.
 Agents do not cause lethal infections ,are not transmissible via
airborne route.
 Agents are pathogens for which immunization or antibiotic
treatment is available.
 Extreme care should be taken with contaminated needles and
sharp lab instruments.
 Class -1 cabinets are used.
 Personal working in BSL -2 labs are expected to take even
greater care to prevent injuries such as cuts and other breaches
of the skin,as well as ingestion and mucous membrane
exposures.
 Personal protective equipment : lab
coats ,gloves ,face protection as needed.
 Protective clothing removed when personnel leave laboratory
area.
Secondary barriers :- BSL -1 barriers plus autoclave for glassware.

17. Standard practices include BSL -1 plus :-
 Appropriate personal protective equipment (PPE) must be worn, including lab coats
and gloves .
 Eye protection and face shields can also be worn ,as needed.
 An autoclave or Appropriate personal protective equipment (PPE) must be worn,
including lab coats and gloves .
 Eye protection and face shields can also be worn ,as needed.
 An autoclave or an alternative method of decontamination is available for proper
disposals .
 This laboratory has self – closing , lockable doors.
 Biohazard warning signs posted outside lab .
 Supervisory staff who have experience working with infectious agents and specific
training for laboratory personnel in handling these agents.
 A sink and eyewash station should be readily available .
 Access to a BSL-2 lab is far more restrictive than a BSL-1 lab.
 Outside personnel ,or those with an increased risk of contamination ,are often
restricted from entering when work is being conducted.
 Primary barriers :- biosafety cabinets or other approved containment devices.
 an alternative method of decontamination is available for proper disposals .

18. BSL -3(special diagnostic service,
research):-
 BSL -3 laboratory typically includes work on microbes that are either indigenous or exotic
,and can cause serious or potentially lethal disease through inhalation .
 Examples of microbes:- Yellow fever ,West Nile virus and bacteria that causes tuberculosis.
 The microbes are so serious that the work is often strictly controlled and registered with the
appropriate government agencies.
Standard practices include BSL -2 lab plus:-
Strictly controlled access to the lab
 Specific training for lab personnel in handling potentially lethal agents .
 Decontaminating all waste
 Changing containmented protective lab coating,decontaminating lab clothing before
laundering .
 Access hands free sink and eyewash are available near the exit.
 Laboratories- under medical surveillance and receive immunisation.

19. Primary barriers :-
 Similar to BSL -2 personnel protective equipment :- respiratory
equipment if risk of infection through inhalation .
 Secondary barriers:- all BSL-2 barriers.
 A self closing set of locking doors with access away from general
building corridors.
 Air handling systems to ensure negative air flow ( air flow into the
lab).
 Air pumped into lab not re circulated in building.

20. BSL 4 ( Dangerous pathogens units):-
 BSL -4 labs are rare.
 However some do exist in a small number of places in the US and around the
world.
 BSL-4 lab consist of work with highly dangerous and exotic microbes
 Agents include- Ebola virus
 Primary hazards:- respiratory exposure to infectious aerosols
. Mucous membrane exposure to infectious Droplets.
Standard practices include BSL-3 plus:-strictly controlled access to the
laboratory.
Decontamination of all materials before existing.
 Personnel are required to change clothing before entering,shower upon
exiting.
 Personnel must wear appropriate personal protective equipment from prior
BSL levels,as well as full body ,air- supplied , positive pressure suit.
 BSL-4 laboratory is extremely isolated often located in a separate building or
in an isolated and restricted zone of the building.

23. References:-
www.labogene.com
www.nuaire.com
www.escoglobal.com