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Biosafety Training. University of Ottawa * Office of Risk Management  Human Resources - Occupational Health Disability & Leave. v0501. Biosafety Outline. Introduction Laboratory Associated Infections Blood-borne Pathogens Classification of Biohazards Infection/Biohazard Control

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biosafety training
Biosafety Training

University of Ottawa

*Office of Risk Management

Human Resources - Occupational Health Disability & Leave

v0501

biosafety outline
Biosafety Outline
  • Introduction
  • Laboratory Associated Infections
  • Blood-borne Pathogens
  • Classification of Biohazards
  • Infection/Biohazard Control
  • Spill Response
  • Biomedical Waste
  • Regulations
what is biosafety
What is Biosafety?
  • Measures employed when handling biohazardous materials to avoid infecting oneself, others or the environment.
  • Achieved through
    • Engineering Controls
    • Administrative Controls
    • Practices and Procedures
    • Personal Protective Equipment
what is a biohazard
What is a Biohazard?

A potential hazard to humans, animals or the environment caused by a biological organism, or by material produced by such an organism

Examples;

  • Viruses, bacteria, fungi, and parasites and their toxins.
  • Blood and body fluids, as well as tissues from humans and animals.
  • Transformed cell lines and certain types of nucleic acids .
who s responsible who are the stakeholders
Who’s Responsible, who are the Stakeholders?

INTERNALLY

  • Vice-President (Research)
  • Committees
  • University Services (ORM, HR, PRS, PS)*
  • Deans, Chairs, Principal Investigators, Employees, Students
  • Manager of Biological Containment Suite

EXTERNALLY

  • Health Canada
  • Canadian Food Inspection Agency
  • Transport Canada
  • Ontario Ministry of Labour
  • Emergency Response Personnel
  • Suppliers & Contractors
  • Community
key services
Key Services
  • Office of Risk Management
    • Training
    • Interface with Regulatory Bodies
    • Biosafety Program
  • certifications
  • training
  • procedures
  • inspections
  • contingency planning
  • accident/incident follow-up
key services7
Key Services
  • HR (Occupational Health, Disability and Leave)
    • Medical surveillance
    • Immunizations
    • Medical Follow-up
    • Interface with Workplace Safety and Insurance Board
why are we concerned about biohazardous materials
Why are we concerned about biohazardous materials?
  • Potential for acquiring a laboratory-associated infection (LAI)
  • Contamination of the environment
  • Contamination of research
  • Public perception*
laboratory associated infections
Laboratory Associated Infections
  • Percutaneous inoculation
  • Inhalation of aerosols
  • Contact of mucous membranes
  • Ingestion

Infection Source

Susceptible Host

  • Immune system
  • Vaccination status
  • Age
  • Cultures and stocks
  • Research animals
  • Specimens
  • Items contaminated with above

Route of Transmission

lai s
LAI’s
  • Only 20% causative or defined event
    • 80% of which are caused by human error
    • 20% are caused by equipment failure
  • Top 4 accidents resulting in infection
    • Spillages & splashes
    • Needle and syringe
    • Sharp object, broken glass
    • Bite or scratch from animals or ectoparasites

http://www.weizmann.ac.il/safety/bio2.html

bloodborne pathogens bbp
Bloodborne Pathogens (BBP)
  • Sources
    • Blood
    • Semen
    • Vaginal Secretions
    • Body Fluids
      • Cerebrospinal
      • Amniotic
      • Synovial
    • Tissue Cultures
    • Organ Cultures
    • Infected Experimental Animals
risk of exposure
Risk of Exposure
  • Pathogen involved
  • Type of body fluid
  • Route of exposure
  • Duration of exposure
  • Volume of blood involved in exposure
  • Concentration of virus at time of exposure
  • PPE worn
issues to consider
Issues to Consider
  • Symptoms
  • Mode of transmission
  • Incubation period
  • Survival outside host
  • Communicability
  • Immunization
  • Prophylaxis / Treatment
if an exposure occurs or the possibility of exposure
If An Exposure Occurs (or the possibility of exposure)
  • Initiate first aid
  • Notify your supervisor / designated person
  • Report to hospital emergency department or University’s Health Services
  • Report incident to OHDL

Occupational Health, Disability and Leave Office telephone ext. 1472 http://www.uottawa.ca/services/hr/frames.html

universal precautions
Universal Precautions
  • Minimum standard of practice for preventing the transmission of BBP

Includes: - Education

- Hand washing

- Wearing protective barriers

- Use safe work practices

If samples cannot be guaranteed non-infective …… treat as infectious!

classification of biohazards
Classification of Biohazards
  • Conventional Agents
  • Unconventional Agents
  • Recombinant DNA
  • Tissue Culture
  • Animal Work
  • Anatomical Specimens

Class D, division 3 of WHMIS (Poisonous and Infectious Material - Biohazardous Infectious Material)

classification of biohazards20
Classification of Biohazards

_

  • As the level  so does ;
  • the risk of the organism to humans, animals, plants and/or the environment
  • the procedural and facility requirements
  • the level of containment required
  • the degree of protection for personnel, the environment and the community.

BSL 4

BSL 3

BSL 2

BSL 1

_

conventional agents
Conventional Agents

Unlikely to cause disease in healthy workers or animals

Rarely cause serious human or animal disease

May cause serious disease

Likely to cause very serious disease

unconventional pathogens
Unconventional Pathogens
  • TSE prion diseases; lethal transmissible neurodegenerative conditions
    • Creutzfeld-Jakob disease, Variant C-J Disease, Mad Cow Disease, Scrapie, Chronic Wasting Disease.
  • Resistant to destruction by procedures that normally inactivate viruses.
  • Contact ORM to assess requirements (containment, procedures, waste disposal, etc.)
recombinant dna
Recombinant DNA
  • Canada: Level of risk depends on source of DNA, vector and host.
  • The Biosafety Committee will assist the investigator in this determination.

Genetic Engineering = in vitro incorporation of genetic material from one cell into another

tissue culture
Tissue Culture
  • Have the potential to contain pathogenic organisms
  • In general;

Human & non-human primate, and mycoplasma-containing cell lines

Level 2

Level 1

Others

A detailed risk assessment should be undertaken when using a new cell line.

animal work
Animal Work
  • Animals can harbour infectious organisms (naturally or introduced)
  • Level dependent on type of work being conducted.
  • Special Animal Care training is required for all personnel working with animals.
  • All work involving animal use must receive prior approval from the Animal Care Committee
anatomical specimens
Anatomical Specimens
  • All specimens should be considered infectious due to potential presence of infectious agents
  • Important to consider the type of specimen
    • blood, organs, tissues
    • Spinal sample, brain tissue
    • From infectious patient
  • In general Level 2 but it depends on the nature of the work.
infection biohazard control
Infection/Biohazard Control

Engineering Controls

Administrative Controls

Practices and Procedures

Personal Protective Equipment

engineering controls
Engineering Controls
  • Technology based, reduce or eliminate exposure to hazards by changes at the source of the hazard.
  • Containment:
    • Primary vs Secondary
    • Containment levels
primary containment
Primary Containment
  • First line of defence.
  • Ensures protection of personnel and immediate environment from exposure to the infectious agent.
  • ‘Protective envelope’ that encapsulates the infectious agent or animal.
    • Petrie dish, vial, stoppered bottle….
    • Biological safety cabinets, glove boxes and animal caging equipment….

Effectiveness of control is based on the integrity of the containment.

secondary containment
Secondary Containment
  • Protects the environment external to the laboratory from exposure.
  • Includes facility design and operational practices.
biosafety containment levels
Biosafety Containment Levels
  • Containment Levels similar to Risk Levels.
  • Biohazards Committee will evaluate the research proposals to ensure adequate containment .
    • Level 1
    • Level 2
    • Level 3
    • Level 4
level 1
Level 1
  • Basic laboratory
  • Requires no special design features
  • Biosafety cabinets are not required and work may be performed on the open bench.
level 2
Level 2
  • Clinical, diagnostic, research and teaching facilities with level 2 agents.
  • Requires a class I or class II biological safety cabinet if any potential for aerosol or splash exists.
  • An emergency plan for handling spills must be developed.
  • Access should be controlled.
slide34
Level 3
  • Specialized design and construction
    • primary barriers to protect the individual
    • secondary barriers to protect the environment
  • All staff must undergo special training on the agents being used, PPE, equipment, waste management as well as practices and procedures above and beyond the scope of this course.
level 4
Level 4
  • Only one level 4 facility in Canada (Canadian Centre for Human and Animal Health in Winnipeg, Man.)
  • Design specifications are extremely stringent, worker is completely isolated from infectious material.
biological safety cabinets
Biological Safety Cabinets
  • Effective means of physical containment for biological agents, especially when aerosols are generated.
  • HEPA filters remove particles (min 0.3 microns) with 99.97% efficiency.
  • There are 3 main classes of cabinets (I, II, III) which provide various levels of protection.
biological safety cabinets37
Biological Safety Cabinets

VS

  • Laminar flow hoods
  • NOT biological safety cabinets
  • Vertical or horizontal laminar flow
  • HEPA filtered air (intake)
  • product protection only
  • Biological Safety Cabinet
  • HEPA filtered laminar air flow and
  • exhaust
  • personnel, environment & often
  • product protection
working safely in a bsc
Working safely in a BSC

Before using the cabinet:

  • Ensure BSC is certified
  • Turn off UV lamp; turn on fluorescent lamp
  • Disinfect work surfaces with appropriate disinfectant
  • Place essential items inside cabinet
  • Allow the blower to run for 5-10 min before work
slide39
Working safely in a BSC

During use:

  • Ensure material and equipment is placed near the back of the hood, especially aerosol-generating equipment. Do not block any vents.
  • Use techniques that reduce splatter and aerosols.
  • General work flow should be from clean to contaminated areas.
  • Minimize movement so as not to impede air flow.
  • Open flame in BSC’s is controversial.
slide41
Working safely in a BSC

After completion of work:

  • Leave blower on at least 5 minutes to purge cabinet
  • Remove and decontaminate equipment and materials
  • Disinfect cabinet surfaces
  • Turn off blower and fluorescent lamp, turn on UV lamp
working safely in a bsc42
Working safely in a BSC

Maintenance:

  • Twice daily - Work surfaces wiped down
  • Weekly - UV lamp should be wiped clean*
  • Monthly - All vertical surfaces wiped down
  • Annually - UV lamp intensity verified.

- Decontamination with formaldehyde gas (ORM)

- Certification (ORM)

administrative controls
Administrative Controls

Program based, information and methods to minimize risk of exposure.

  • Risk assessment
  • Medical Surveillance
  • Training/Education
  • Resources
  • Inspections
  • Signs & Labeling
administrative controls44
Administrative Controls

Risk Assessment

  • Will determine type of containment, procedures, and safety equipment required
  • Responsibility of users, additional assistance is available from ORM
  • Consider areas such as; experimental design, procedures to be employed and personal experience/knowledge, etc.

*

http://www.hc-sc.gc.ca/pphb-dgspsp/ols-bsl/lbg-ldmbl/pdf/lbg-3e-draft.pdf

administrative controls45
Administrative Controls

Risk Assessment: Know your Agent

  • Know and understand the various characteristics of the agent(s) you are working with.
  • This information is available from;
    • MSDS’s
    • Suppliers or manufacturers
  • Example
administrative controls46
Administrative Controls

Medical Surveillance

Training & Education

  • Lab specific policies and procedures
  • Biosafety training

Resources

  • ORM web site, Biosafety page
  • Faculty web sites
  • Biosafety Manual
  • Training Videos
administrative controls47
Administrative Controls

Inspections

  • Routine self-inspections
  • Biosafety Inspection Checklist available on-line
  • In addition, ORM, EHSOs and OH&S will inspect labs to ensure compliance with regulations/ guidelines and provide feedback.
administrative controls48
Administrative Controls

Signs & Labeling

  • Biohazard warning signs must be posted on doors to rooms where biohazardous materials are used.
  • Biohazard labels should be placed on containers, equipment and storage units used with biological agents.
practices and procedures
Practices and Procedures
  • General Safety Guidelines
  • Good Microbiological Practice
  • Handwashing
  • Specific Procedures
    • Centrifuges
    • Needles & Syringes and other sharps
    • Pipettes
    • Blenders, Grinders, Sonicators & Lyophilizers
    • Inoculation Loops
    • Cryostats
general laboratory safety guidelines
General Laboratory Safety Guidelines
  • Mostly common sense, but you must understand the hazards you face in the laboratory and be adequately trained to deal with them.
  • Basic must knows for all labs.
  • Examples?

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good microbiological practice gmp
Good Microbiological Practice (GMP)
  • Basic code of practice that should be applied to all types of work involving microorganisms
  • Objectives
    • prevent contamination of laboratory workers and the environment
    • prevent contamination of the experiment/samples
  • Application of aseptic technique, minimization of aerosols, contamination control, personal protection, emergency response
handwashing
Handwashing
  • One of the single effective means of preventing infections
  • IF done properly and frequently
  • When to wash?
  • Before starting any manipulations
  • Before leaving the lab
  • When hands are obviously soiled
  • Before and after completing any task in a BSC
  • Every time gloves are removed
  • Before contact with one’s face or mouth
  • At the end of the day
safe use of centrifuges
Safe use of Centrifuges
  • Before use
    • Stress lines? Overfilled? Balanced?
    • Caps or stoppers properly in place?
    • Run conditions achieved?
  • Use sealable buckets (safety cups) or sealed rotors
  • After run
    • Centrifuge completely stopped?
    • Spills or leaks?
    • Allow aerosols to settle (30 min) or open in a BSC.
needles and syringes
Avoid use whenever possible

Use a BSC for all operations with infectious material

Fill syringes carefully

Shield needles when withdrawing from stoppers

Do not bend, shear or recap needles.

Dispose of all used needles/syringes in yellow sharps containers

Needles and Syringes
pipettes
Pipettes
  • Mouth pipetting is prohibited.
  • All biohazardous materials should be pipetted in BSC’s.
  • Never force fluids out, use ‘to deliver’ pipettes.
  • To avoid splashes, allow discharge to run down dispense the receiving container wall.
  • Never mix material by suction and expulsion.
  • Reusable pipettes should be placed horizontally in a disinfectant filled pan.
blenders grinders sonicators and lyophilizers
Blenders, Grinders, Sonicators and Lyophilizers
  • Operate in a BSC whenever possible. Allow aerosols to settle for 5 minutes before opening.
  • Safety Blender
    • Do not use glass blender jars
    • Decontaminate immediately after use
  • Lyophilizers
    • Use glassware designed for vacuum work, ensure there is no damage before using.
    • All surfaces should be disinfected after use
    • Use vapour traps whenever possible.
inoculation loops
Inoculation Loops
  • Sterilization in an open flame may create aerosols which may contain viable microorganisms.
  • Shorter handles minimize vibrations.
  • Disposable plastic loops are good alternatives.
cryostats
Cryostats
  • Wear gloves during preparation of frozen sections and heavy gloves when accessing the cryostat.
  • Decontaminate frequently (70% Ethanol or specific to agent)
  • Guards and wheel locks!
personal protective equipment
Personal Protective Equipment
  • PPE can become an important line of defence (last line of defense).
  • Responsibility of both the user and the supervisor to ensure that PPE is worn
slide60
PPE
  • Criteria for consideration
    • Routes of exposure that need to be blocked
    • Degree of protection offered
    • Ease of use
  • Only effective if correctly selected, fitted, used and cared for, and the individual is trained
  • Ensure PPE is removed before leaving the lab.
slide61
PPE

Footwear

  • Closed toed shoes should always be worn. Booties are worn in some higher containment labs and animal facilities.

Lab Coats/Gowns

  • Long-sleeved, knee length with snaps
  • Elastic cuffs *
  • Back-closing gowns *
  • Periodic cleaning required
slide62
PPE

Gloves

  • Latex, nitrile & vinyl for work with biological agents
  • Exam gloves should not be reused, change frequently. Utility gloves can be disinfected and reused if they show no sign of degradation.
  • Consider tensile characteristics, length of cuff
  • Double gloving
  • ORM can provide assistance finding an alternative for people with allergies.
  • Gloves are not to be worn in public places
slide63
PPE

Eye & Face Production

  • Goggles, safety glasses to protect the eyes
  • Full face shield to protect facial skin.

Respirators

  • Only personnel who have been fit-tested and trained should wear respirators.
spills
Spills
  • Spill response will vary depending on:
    • What was spilled?
    • How much was spilled?
    • Where was the spill?
    • What is the potential for release to the environment?
  • Spills should be cleaned up immediately (unless an aerosol was generated), to ensure proper decontamination.
  • Ensure appropriate PPE is worn and clean-up equipment is readily available.
spills general clean up
Spills – General Clean-up
  • Cover spill area with absorbent material
  • Soak the spill area with an appropriate disinfectant (i.e. 10% bleach)
  • Pour disinfectant from the outside surface of the absorbent material towards the inside
  • Ensure any broken glass is picked up (with forceps!) and placed in a sharps container
  • Leave on for 20 to 30 minutes
  • Wipe up with absorbent material
  • Waste should be disposed in appropriate biohazard bags and where possible autoclaved
spills special cases
Spills– Special Cases
  • Within a Centrifuge
  • Within a BSC
  • Open Areas (lab, during transport)
  • The spill response plan template is available at http://www.uottawa.ca/services/ehss/SPILLRESPONSEPLAN.pdf
spills67
Spills
  • All users of biological materials should be familiar with the spill clean-up procedures.
  • All spills are to be reported ASAP to the lab supervisor and ORM.
  • Additional assistance is available from:
    • ORM x 5892
    • Your departmental safety officer
    • ERT x 5411 (through Protection)
decontamination disinfection and sterilization
Decontamination, Disinfection and Sterilization
  • Decontamination: Free of contamination, the destruction of microorganisms to a lower level such that it removes danger of infection to individuals.
  • Sterilization: The complete destruction of all viable microorganisms.
  • Disinfection: Use of agents (physical or chemical) to destroy harmful organisms on inanimate objects (not necessarily all organisms)
decontamination physical
Decontamination: Physical
  • Heat:
    • Autoclaving (most practical and recommended)
    • Incineration (for disposal of sharps and tissues)
  • Irradiation:
    • UV light (wavelength of 253 nm is germicidal)
    • Gamma (disrupts DNA and RNA)
  • Filtration
    • HEPA (biological safety cabinets, ventilation)
autoclaves
Autoclaves

Items that CAN be autoclaved:

  • Cultures and stocks of infectious material
  • Culture dishes and related devices
  • Discarded live and attenuated vaccines
  • Contaminated solid items (petrie dishes, eppendorf tips, pipettes, gloves, paper towels)

Items that CAN NOT be autoclaved:

  • Chemical, chemotherapeutic or radioactive waste
  • Bleach
  • Certain kinds of plastics
  • Sharps (not at the University of Ottawa)
autoclaves71
Autoclaves

Preparation of waste:

  • Use only approved autoclave bags.
  • Do not overfill autoclave bags
  • Separate material for re-use from that which will be disposed and dry from liquid material .
  • If outside of bag is contaminated, double bag.
  • All flasks containing biological material should be capped with aluminum foil.
  • Ensure items are labeled with contact information.
safe use of autoclaves
Safe Use of Autoclaves
  • Many autoclaves are now run by dedicated staff, however, if you are operating an autoclave;
    • Learn how to use!
    • Ensure PPE is worn
    • Recognize acceptable material and packaging
    • Proper loading and unloading

All users/operators must fill out the Autoclave User Questionnaire and receive training!

decontamination chemical
Decontamination: Chemical
  • Generally for disinfection rather than sterilization
  • Choice depends on;
    • Type of material to be disinfected
    • Organic load
    • Chemical characteristics
  • Most common are chlorine compounds and alcohols (broad range)
disinfection what to use for my organism
Disinfection: What to use for my organism?

Bacteria

Vegetative bacteria(E.coli, Staph)

  • 2% domestic bleach
  • 75% Ethanol
  • Quaternary ammonia
  • 6% formulated Hydrogen peroxide*

Mycobacteria and fungi

  • 10% domestic bleach
  • 75% Ethanol
  • Phenolic compounds
  • 6% formulated Hydrogen peroxide*

Spore forming bacteria(Bacillus)

  • 10% domestic bleach
  • Gluteraldehyde
  • Formaldehyde
  • 6% formulated Hydrogen peroxide*

Viruses

Enveloped (HIV, Herpes)

  • 2% domestic bleach
  • 75% Ethanol
  • Quaternary ammonia
  • 6% formulated Hydrogen peroxide*

Non enveloped (Hepatitis, Adenovirus)

  • 10% domestic bleach
  • 6% formulated Hydrogen peroxide*
  • Gluteraldehyde
  • Formaldehyde
waste management
Waste Management

Biomedical waste

Discarded biological material from teaching, clinical and research laboratories and operations. Biomedical waste includes but is not limited to;

  • Animal waste
  • Biological laboratory waste
  • Human anatomical waste
  • Human blood and body fluid waste
  • Sharps
waste management76
Waste Management
  • All biological waste should be decontaminated prior to disposal (including level 1 agents).
  • Treated waste is no longer considered ‘biomedical’ (i.e. microbiological waste, blood and bodily fluid waste) and can be disposed in the regular waste stream.
  • Any waste that cannot be treated (i.e. sharps, carcasses, tissues and body parts) remains biomedical waste and must be incinerated off site.
waste disposal
Waste Disposal

Biomedical Waste (untreated)

waste disposal78
Waste Disposal

Biomedical Waste (treated)

*

In compliance with Sewer use by-laws

With H2O 1:10

special waste
Special Waste
  • EtBr
  • Toxins
  • Recombinant DNA
  • Contact ORM
transportation
Transportation
  • Important Considerations:
    • does material need to be transported at all
    • packaging requirements
    • means and route of transportation
    • regulatory requirements
  • Between lab transfers - 4 sided cart, sealed primary container, secondary container, low traffic route.
  • Off Campus transfers – consult ORM
shipping receiving
Shipping & Receiving
  • Transportation of Dangerous Goods Act: Class 6.2 of (Infectious Substances)
  • International Air Transport Association
  • HC/CFIA restrictions
  • Ensure;
    • Proper classification
    • Proper packaging
    • Proper labeling
    • Proper documentation
    • Import/Export Permits
purchasing
Purchasing
  • Importation permits required by Health Canada or CFIA for certain agents
  • US restrictions
  • Ensure you meet all criteria and have all pertinent documentation
security
Security
  • Controlled access
    • physical barriers (structural design, departmental design, key/card access, etc.)
    • psychological barriers (obvious presence of identifiable security personnel, security culture, use of monitoring tools)
    • monitoring activities (security patrols, departmental monitoring, key control program)
    • personnel clearance
inventory
Inventory
  • What material is presently being used and/or stored in the lab
    • Location
    • Expiry date
    • How much, use log
    • MSDS’s
the bottom line
The Bottom Line
  • If you are notcareful and diligent with biological agents you risk:
    • Infecting yourself, others or the environment
    • Contaminating your research
    • Having Health Canada, Canadian Food Inspection Agency, Ministry of the Environment or Transport Canada after you
conclusions
Conclusions
  • Biosafety - ensuring that individuals and the environment are not infected
  • Biohazards - microorganisms, blood and body fluids, tissues and tissue culture
  • Everyone within the University community is responsible
  • With proper knowledge, planning and care, a biological exposure is avoidable.
biosafety website orientation
Biosafety Website Orientation

http://www.uottawa.ca/services/ehss/biosafety.htm

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