INDUSTRIAL SAFETY & HEALTH

CKB 30103
BIOHAZARD
 OBJECTIVES
At the end of this chapter, you will be
able to;
• Define the term of biohazard
• Able to classify the type of biohazards and
their risk level.
• Define the term of biosafety and
understand the biosafety level.
• Understand the control of biohazard and
various type of decontamination methods.
 Some unfortunate examples of disease outbreaks in research labs:

1950‐1976:

• •A survey of 5000 labs showed 3921 cases of disease outbreaks

• •Most commonly reported were: Hepatitis, tuberculosis, typhoid, brucellosis, rabbit
• fever

More recently (since 2003):

• •SARS infects researchers in a lab in Singapore
• •A US and a russian scientist are infected by Ebola. One survives, one dies.
• •Polio virus escapes from two idian labs
• •Scientists from Boston University contract rabbit fever (a serious bacterial disease)
• •Anthrax exposure in a Huston lab due to aerosols leaked inside an unshielded
• centrifuge
 Definition

Biohazard

Biohazard are infectious agents or

biologically infectious material, which
presents or may present a risk to the
well being of a person or the
community
 Definition

Biohazard might also be defined as

the infectious agents presenting a
risk of death, injury or illness to
employees

…. The Biohazard Technology Texas Collaborative Team

Categories of biohazard
Human, animal and plant pathogen [Bacteria,
Fungi, Viruses, Parasites, Rickettsiae, Chlamydiae, Toxins Bacteria]

Human & animal blood, blood product,

tissues and body fluids
Cultured cells & potentially infectious
agents [Prion-proteinaceous product]
Allergens [dust]
Recombinant DNA products [GMO –genetically
Modified Organisms]

Clinical, necropsy & surgical specimens

(tissues, fluids)
Snake venom
Virus
Fungi
Baceria
Source of Biohazard
• Human Waste - All human blood, tissues, body fluids,
secretion, excretions, organs, and cadavers. All items
contaminated with human waste are also considered to
be infectious biohazard waste.

• Sharps Waste - All needles, syringes (with or without

the attached needle), pasteur pipettes, scalpel blades,
needles with attached tubing, suture needles, and
slides, cover slips and plastic ware.

• Microbiological Waste - All materials containing or

in contact with cultures of microbiological organisms and
all that were cultured or items contaminated by
specimens.
Source of Biohazard cont……
• Biological Materials Waste - All discarded
vaccines, immune globulins, plasma, albumin blood or
tissue fractionation products, enzyme preparations, etc.

• Animal Waste - All animal carcasses, tissues, blood,

body fluids, and all items grossly contaminated with
these items.

• Animal Pathogen Contaminated Waste

- All bedding and other materials contaminated with
blood, excreta or secretions of animals infected with
transmissible human or animal pathogens.
 Risk Groups
• Based on transmissibility,
invasiveness, virulence and lethality
of the specific pathogen
• RG1 – not associated with disease
• RG2 – associated with disease that is
rarely serious or for which there is
treatment
• RG3 – serious or lethal human
disease with treatments
• RG4 – serious or lethal human
disease with no treatment options
 Definition

Biosafety:
The containment principles, technologies
and practices that are implemented to
prevent the unintentional exposure to
pathogens and toxins, or their accidental
release
 Biosafety Level

• A biosafety level is the level of the biocontainment

precautions required to isolate dangerous biological
agents in an enclosed facility. The levels of
containment range from the lowest biosafety level 1
to the highest at level 4.

• Correlates to Risk Groups

• Describes containment practices, equipment, and

facility design features recommended for safe
handling of these organisms
 Biosafety Level

• BSL-1: agents are not known to

cause disease
• BSL-2: agents are associated with
human disease
• BSL-3: agents are associated with
human disease and are potentially
transmitted as aerosols
• BSL-4: agents of life threatening
nature
 Biosafety Level 1
• Class 1 agents are not associated with disease in
healthy adult humans

• However, increased precautions may be

appropriate when using these agents for
transfection since foreign genes can be delivered
even though disease is not caused
• Some examples include:
• E. coli nonpathogenic laboratory strains
• Saccharomyces cerevisiae
• Agrobacterium tumefaciens
• Baculovirus
• Duck hepatitis B virus
 Risk Group 1 Agents

• E.coli K-12
• Transgenic Plants
• Plasmids
• Fungi
• Mold
• Yeast
 Biosafety Level-1
Concepts of Biosafety
Biosafety Level-1 (BSL-1)
• Well characterized agents
• Agents not known to cause disease (in
healthy human adults; now healthy immunocompetent
adults)
• Prophylactic treatment available
• Open bench procedures
• Animals in open cage system or open
environment (outdoors)
• Good laboratory practices
 BSL-1 Practices
• Bench-top work allowed
• Daily Decontamination
• Manual pipetting
• Required Handwashing
• Red bag waste
• Bio cabinet not required
(unless creating aerosols)
• 2˚ containment
 Biosafety Level 2
• Class 2 agents are associated with
human disease which is rarely
serious and for which preventative or
therapeutic interventions are often
available.
• Some examples would include:
• E. coli, pathogenic strains
• Adenovirus
• Herpes simplex virus
• Chicken pox
• Moloney murine retrovirus
 Risk Group 2 Agents

• Human or Primate Cells

• Herpes Simplex Virus

• Replication Incompetent
Attenuated Human
Immunodeficiency Virus

• Patient specimens
 BSL-2 Practices
Concepts of Biosafety

Practices & Procedures

• Agents associated w/ human disease
• Treatment for disease available
• Agent poses moderate hazard to
personnel and environment
• Direct contact or exposure
• Percutaneous exposure
• Scratch, Puncture, Needle stick
• Mucus membrane exposure
• Eyes, Mouth, open cut
 BSL-2 Practices

• Limited access to lab when

work in progress
• Daily decontamination
• Mechanical pipetting
• Labcoat, safety glasses and
gloves required
• Red bag & sharps containers
required
 BSL-2 Practices (con’t)
• Biohazard Sign posted at
entrance to lab
• Label all equipment
(incubators, freezers,
etc.)
• TC room – negative air
flow
• Documented training
• Baseline serology or pre-
vaccination may be
required
 Biosafety Level 3
• Class 3 agents are associated with serious or
lethal human diseases for which preventative
or therapeutic interventions may be available
(high individual risk but low community risk)
• Some examples would include:
• Francisella tularensis
• Human immunodeficiency virus
• Histoplasma capsulatum
• Mycobacterium tuberculosis
• Aerosol is a common route of transmission,
and increases the risk potential for these
agents
 Risk Group 3 Agents

• Human
Immunodeficiency
Virus
• Mycobacterium
tuberculosis
• Coxiella burnetii
 Biosafety Level 3
Working in High Containment

Biosafety Level-3 (BSL-3)

• Indigenous or exotic agents
• Aerosol transmission
• Serious health effects
• Treatment may or may not
exist
 BSL-3 Practices
• Public access NOT permitted
• Daily decontamination after spill
and upon completion of
experiment
• Autoclave required and waste is
disposed at the end of day
• Required foot activated hand
washing sink and controls
• No sharps unless absolutely
necessary
 BSL-3 Practices (con’t)

• Aerosol minimization procedures

required
• Wrap around disposable clothing is
required. Specialized equipment may be
required depending upon procedures
• Biohazard Signs and labels posted
• Air flow from low hazard to high hazard
“Pressure Mapping”
 BSL-3 Practices (con’t)
• Bench top work not permitted
• Documented training and personnel
competency certification (for BSL-3
procedures)
• Baseline serology
• Spills – report immediately and treat
accordingly
• Vaccinations/post exposure protocols
and SOP’s, Biosafety Manual, Biosafety
Officer
BSL-3
 Biosafety Level 4
• Class 4 agents are likely to cause serious
or lethal human disease for which
preventive or therapeutic interventions
are not usually available (high individual
risk and high community risk)
• Some examples would include:
• Ebola virus
• Herpes B virus (Cercopithecine)
• Lassa fever virus
 Risk Group 4 Agent

• Lassa Fever Virus

• Ebola
Hemmorrhagic
Fever Virus
• Marburg Virus
• Herpes B Virus
 Biosafety Level-4
Working in High Containment

Biosafety Level-4
• Builds on BSL-3
• Maximum containment facilities
• Pressurized Containment Suite
• BSL-3 + Class III Biosafety Cabinet
• Chemical decontamination showers
• Liquid effluent collection /
decontamination
• No BSL-4 labs exist in Malaysia
 Biosafety Concepts
Working in High Containment
Biosafety Level-4 (BSL-4)
• Dangerous/exotic agents
• Life threatening disease
• Aerosol transmission
• Agents of unknown risk
of transmission or health affects
• No known treatment
Route of transmission

• Ingestion
• Skin contact
• Respiratory
• Introduction by arthropods/mammal into
lymphatic/blood system
Route of Entry - Ingestion
 A variety of organisms that are around us, especially in working
area are enteric pathogens which use ingestion as the primary
route of infection (intestinal parasites, Salmonella, agents of
infectious hepatitis, polio virus, and enteropathogenic E coli
strains).
 Infection by these organisms generally occurs in the following
ways:
 Direct ingestion
Such as when working in the laboratories which requires
working with organisms, or direct ingestion of the culture
when performing mouth pipetting
 "Hand to mouth" infection
Infectious materials are transmitted indirectly by the hand to
the oral cavity.
 Activities such as smoking, eating, and drinking are therefore
prohibited in work areas especially in the laboratories. Frequent
hand washing with germicidal soap between activities is highly
recommended.
Route of Entry - Skin contact

• Contact can be avoided by limiting the use of needles

and syringes and by using non-breakable containers
whenever possible.

• Workers' hands must never come into direct contact with

infectious agents. Therefore, gloves should be worn and
discarded appropriately before handling other equipment
or objects.
Route of Entry - Respiratory
A variety of agents infect by the respiratory
route. Aerosol generation and dissemination can be
reduced by the following:
• Properly operating laminar-flow biological safety
cabinets for protection against immediately
generated aerosols
• Thorough decontamination of work surfaces
before and after work and following
spills of bio-hazardous material. This method is
particularly effective in preventing secondary
aerosols generated by agents resistant to drying
• Use of absorbent materials on immediate work
surfaces to contain splashes and drips
Preventive Method

• Wear appropriate PPE

• Never discard syringes, needles or any
apparatus used in testing involve
microorganism
• Employ good, sterile techniques when
open any tubes, bottles and culture
plates-minimize contamination
• Storage vessels containing biohazard
agents must be labeled to provide
identification of their content
 Summary
Risk BSL Level Examples Practices Facilities,
Group Equipment

1 Basic Basic Teaching, Good None required;

BSL 1 Research Microbiological open benchwork,
Techniques (GMT) directional
airflow
2 Basic Primary health Level 1 plus Open bench plus
BSL 2 services, diagnostic, protective Biological Safety
teaching, public clothing, Cabinet (BSC)
health biohazard signage for aerosols

3 Containment Special diagnostic, Level 2 plus BSC and/or other

BSL 3 research special clothing, primary
controlled access containment
devices for all
activities
4 Maximum Dangerous pathogen Level 3 plus Class III BSC or
Containment unit airlock entry, positive
BSL 3 shower exit, pressure suits,
special waste double door
disposal autoclave,
filtered air
 Biosafety Cabinets (BSCs)

• Provide product, personal and

environmental protection.
• Various classes of BSCs are available
• Amount of air exhausted
• Amount of employee protection
Biosafety Cabinets (BSCs)
 Biosafety Cabinets (BSCs)
• Sterile air is exhausted from BSCs
through a HEPA filter.
• HEPA filter can trap particles to 0.3 microns
• Chemical vapors and fumes pass through
HEPA filters
 Biosafety Cabinets (BSCs)

Helpful Hints
• Enter straight into the cabinet,
avoid sweeping motions
• Don’t place materials on the grill
• Keep discard pan or bag inside the
cabinet
• Decon materials before removal
from the cabinet
 Decontamination

• Sterilization: destroy all microbial

life, including spores
• Disinfection: destroy a majority of
microbial life, but not necessarily
spores
• Antiseptic: germistat used on skin to
inhibit growth of microorganisms
 Decontamination

Methods
• Heat: steam heat, dry heat,
incineration
• Chemical: bleach, ethanol, hydrogen
peroxide, ethylene oxide,
paraformaldehyde
• Radiation
Sterilization and Disinfection
Sterilization Disinfection

Process that kill / remove all Process intended to kill/remove

types of microbes including pathogens with exception of
resistant spores. spores (spores can only be
removed by sterilization)

Types of sterilization methods: Types of disinfectants:

•Moist/dry heat (steam) •Formalin

•Chemical e.g Ethylene Oxide •Formaldehyde
•Ionizing radiation e.g gamma •Lysol
rays •Alcohol etc
Comparison between different methods of industrial
sterilization
Criteria under Gamma Ethylene Steam
consideration

Material of construction Some plastics discolor Most material ok Some plastics

Some fibre degrade melt

Product packaging No restriction Permeable material Permeable

No stress on seals Seal must stand material
Non permeable ok pressure & Vacuum Expansion
package
Parameters needing Time [EtO] Vacuum,Pressure
control during process Vacuum,Pressure Temperature,RH
Temperature,RH Time
Time
Reliability of process Excellent Good Excellent

Post sterilisation Dosimetric release Biological indicator Parametric release

testing
Comparison between different methods of industrial
sterilization cont……
Consideration Gamma Ethylene Steam

Quarantine period No quarantine 3-14 days 0 – 7 days

required

Post sterilisation None required Aerate to remove Dry product

treatment residues

Economics Good on low & Good on low & Good on low &
high volumes high volumes high volumes
High capital High capital Inexpensive
investment investment operations
Validation Material Complex Simple
qualification performance runs
lengthy
Audit/Requalification Quarterly Yearly Yearly

Ranking / score/10 1 (5/10) 2 (2/10) 3 (1/10)

 Thank you
End of Chapter