FDS

Biosafety TrainingShulin Chen Lab

Developed and posted 11/14/08

http://www.bio-safety.wsu.edu/biosafety/

Outline

• Introduction

• Biosafety Levels Practices Facilities

• Biosafety Cabinet

• Case Study

• Resources

• Regulations

Biosafety Introduction

Practicing Science Safely Means That Before Any Activity...

YOU KNOW the risks YOU KNOW the worst

things that could happen YOU KNOW what to do if

they should happen YOU KNOW AND USE the

prudent practices, protective facilities, and protective equipment needed to mitigate the risks

Biosafety Levels – Know the Risks• BSL 1: Material not known to consistently cause disease in

healthy adults.

• BSL 2: Associated with human disease. Hazard is from percutaneous injury, ingestion, or mucous membrane exposure. Some agents with environmental or agricultural impact.

• BSL 3: Indigenous or exotic agents with potential for aerosol transmission; disease may have serious or lethal consequences.

• BSL 4: Dangerous/exotic agents which pose a high risk of life-threatening disease, aerosol-transmitted lab infections or related agents with unknown risk of transmission

Appropriate Biosafety LevelAgent Risk Group + Risk Assessment =

Appropriate Biosafety Level

Agent Risk Group Resource:http://www.absa.org/riskgroups/index.html

Risk Group Examples:

RG-1E. coli (standard host vector systems)Most plant pathogense.g., Aschochyta spp.Adenovirus type 1-4, most human cell lines

RG-2Klebsiella, Listeria, E. coli O157, Human Adenovirus, Candida spp. Giardia lamblia, Some human cell lines HEK-293 HeLa (characterized agents)

RG-3Mycobacterium tuberculosis, M. bovis, Coccidioides immitis, HIV, R. richettsii

A Complete Risk Assessment includes• Agent

Characterization (Risk Group RG)

• Personnel Factors (experience)

• Work Activity Factors

• Environmental Factors

• Equipment Factors

• Risk Consequences

• Probability Profile

Risk Assessment• Risk of Activity – same agent can have

different containment levels:

Procedures that produce aerosols have higher risk

Procedures using needles or other sharps have higher risk

Handling blood, serum or tissue samples may have lower risk

Purified cultures or cell concentrates may have higher risk

Large volumes (>10 L) have higher risk

Protection Should Match the Risk: What is the Protection Should Match the Risk: What is the Expected Route of Infection?Expected Route of Infection?

AGENT / LAI**LAI= Laboratory Acquired Infections

PRIMARY HAZARDS

Klebsiella spp.

LAI’s have been reported

Direct contact of mucous membranes with contaminated objects, inhalation of infectious aerosols; accidental parenteral inoculation; ingestion

Staphylococcus epidermidis Direct contact of mucous membranes, accidental parenteral inoculation.

AGENT Pathogenicity/Epidemiology

Klebsiella spp. Pathogenicity: Frequent cause of nosocomial urinary and pulmonary infections; wound infections, secondary infection in lunch of patients with chronic pulonary disease, enteric pathogenicity, (enterotoxin).

Epidemiology: Worldwide distribution. 2.3 of all infection due to Klebsiella spp. Are hospital-acquired: causes 3% of all acute bacterial pneumonia; common source of nosocomial outbreaks.

Staphylococcus epidermidis Pathogenicity: Generally considered to be part of normal Human flora but has been documented as a pathogen in numberous cases of bacteremia, surgial wounds, urinary tract and ophthalmologic infections especially in immunocompromised hosts.

Epidemiology: World wide distribution. Associated with nosocomial infections.

Know What to Do

• What Constitutes a Biological Exposure Risk?

• What to do When There has Been a Potential Exposure?

• How to Clean up a Biological Spill?

• When Should you Seek Assistance With a Spill Clean up?

What Constitutes a Biological Exposure Risk?

• Breach in Primary or Secondary Containment

• What is Primary Containment?

• What is Secondary Containment?

What to do when there is a breach?

Tell your supervisor and the Biosafety Manager for WSU. Fill out an incident report, and all other paper work as required.

What to do when there is a near miss?

Fill out an incident report.

Primary Containment

• Lab practices – standard lab practice, limited access, biohazard warning sign, sharps/needle precautions, SOPs, decontamination, waste.

• Safety equipment – biosafety cabinets (BSC), sharps containers, sealed rotors.

• Personal protective equipment (PPE) – lab coat, gloves, goggles, respirators.

Establishing Precautions to Minimize Risk

Primary containment

Practices/Equipment

PPERisk A

ssessment

Immunization

Surveillance

Secondary Containment - Facilities

• Facility provides containment through traditional construction BSL-1 & BSL-2 Best practices HVAC provides negative

pressure in BSL-2 labs. Depending on risk assessment this may be required.

• Facility provides containment through special design features BSL-3 Anterooms Double doors Alarmed redundant HVAC systems

Possible Biological Release Scenarios

Is This a Release?

1. Open a centrifuged tube at the open bench that contains Klebsiella spp.?

2. Using a wire transfer loop and a bunsen burner pick a Klebsiella colony and streak for isolation work in the BSC.

Is This a Release?

3. Transfer liquid cultures with a transfer pipette to produce BSL-2 organism serial dilutions. Work performed at the open bench.

Answers to Release Scenarios1. Open a centrifuge tube of Klebsiella at open bench

• Centrifugation causes the pressure inside the centrifuge tube to be altered from the ambient pressure. When opening the lid aerosols are formed. Performing this action at the open bench exposes you and your coworkers to potentially infectious aerosols

2. Bunsen burner in the BSC

• Bunsen burners cause air turbulence in the Biosafety Cabinet which may lead to escape of bio-aerosols and subsequent exposure to infectious aerosols to the cabinet worker and others in the laboratory.

Answers to Release Scenarios

3. Infectious liquid transfer at bench

• Any manipulation of infectious liquids potentially creates aerosols. To perform manipulations of infectious liquids safely this work should be done in the Biosafety Cabinet

Summary

In all three of these case studies there has been a potential release of infectious aerosols.

NOTE: The following slide shows a picture of aerosol generation when expelling a liquid from a pipette.

Spill Clean up of Biological AgentsSurface Contamination: Alert co-workers Define/isolate contaminated area Put on appropriate PPE (personal protective equip.)

to include gloves, lab coat and face shield (if appropriate)

Remove/glass/glass shards with forceps or scoop Apply absorbent towels to spill – Do NOT apply

disinfectant directly to the spill as this may aerosolize the agent

Apply disinfectant to towel surface Allow adequate contact time (generally 20

minutes) Remove towels, mop up; clean with alcohol or

soap/water or other agent as appropriate. Dispose of materials in biohazardous waste Notify lab instructor

Spill in a BSC• If the spill of an infectious agent was enough to create

puddles or liquid in the drain pan then the following procedure should be followed: a. Leave the cabinet running and close the view

screen for about 5 minutes. This will allow aerosols to settle before starting cleanup.

b. The drain pan should be flooded with appropriate disinfectant. Leave the disinfectant in the pan for required contact time, longer if the spill involved a high organic load and 10% bleach is used. The disinfectant then needs to be drained out and the surfaces thoroughly cleaned with water to prevent corrosion.

Spill Clean up of Biological AgentsPersonal Exposure:• Clean exposed surface with soap/water,

(1 minute), eyewash (eyes) 15-20 min.,

or rinse mouth 3x’s with water

• Apply first aid and treat as an emergency

• Notify lab instructor – fill out incident report and other forms as requested

• If appropriate report to medical clinic for treatment/counseling

Major Spills

• This is a spill of a potentially biohazardous material that will take more than 30 minutes to clean up (not including the agent deactivation period of 10-60 minutes depending on the biological agent and disinfectant in use)

Call 911 for assistance

General Guidelines for Surface Decontamination AGENT DISINFECTANT INACTIVATION TIME

Recombinant DNA 10% Bleach 20 minutes

Bacterial Spores >6% Hydrogen Peroxide 30 minutes

Vegetative Bacteria 10% Bleach 20 minutes

Viruses & Viroids 10% Bleach 20 minutes

Fungi 10% Bleach 20 minutes

Feline Parvovirus 20% Bleach 30 minutes

Free Living Cryptosporidium

70% Ethanol 10 minutes

Most Parasites 10% bleach 30 minutes

Prions 50% bleach 60 minutes

Practical Disinfectants for use in Recombinant DNA Research NE=Not Effective, b=variable results dependent on virus

Reference: NIH Guidelines for working with Recombinant DNA Lab Safety Monograph (Appendix D updated) LiquidDisinfectants

Practical Requirements Inactivates Important Characteristics

Use/Dilution Contact time minutes

Vegetative Bacteria

Lipo-viruses

NonlipidViruses

Bacterial Spores

EffectiveShelf life >

1week

Effective for Surface Decontamination

LiquidsFor

Discard

Category Lipo virus Broad Spectrum

Quaternary Ammonia Compounds

0.1-2.0% 10 NE + + + +

Phenolic Compounds

1.0-5.0% 10 NE + + b + +

Chlorine Compounds

500ppm(a) 10 30 + + + + + +

Iodophor 25-1600ppm(a)

10 30 + + + + + +

Alcohol, Ethyl 70-85% 10 NE + + b + +

Alcohol, isopropyl

70-85% 10 NE + + b + +

Formaldehyde 0.2-8.0% 10 30 + + + + + +

Glutaraldehyde 2.0% 10 30 + + + + + +

Know and Use

• prudent practices,

• protective facilities, and

• protective equipment

needed to mitigate risks

Recommended Biosafety Containment & Practices – Infectious Agents

BSL Agents Practices

Primary Containment

Safety Equipment

Primary Barriers

Facilities

(Secondary Barriers)

1 Not known to consistently cause disease in healthy adults

• Wash hands after handling viable materials

• No smoking, eating, handling contact lenses, applying cosmetics

• No mouth pipetting

• Minimize aerosols& splashes

• Decontaminate work surfaces daily

• Do NOT wear gloves out of lab

• Remove gloves before touching door handles phones etc.

None required •Doors

•Sink required

•Work surfaces easily cleaned & impervious to water

•Sturdy furniture

Recommended Biosafety Containment & Practices – Infectious Agents

BSL Agents Practices

(Primary Containment)

Safety Equipment (Primary Barriers)

Facilities

(Secondary Barriers)

2 Associated with human disease, hazard = percutan-eous injury, inges-tion, mucous mem-brane exposure

BSL-1 practice plus:

•Limited accessLab entry and exitpolicies.

•Biohazard warning signs

at entrance of lab

•“Sharps” precautions

•Decontaminate stock cultures prior to disposal

•BSM - SOP

•Report Spills & Accidents

•Leak-proof containers for transport

•PPE required (as approp.)

•Documented training

Primary barriers = BSC I or II or other physical barriers, e.g. splash shields, covered centrifuges etc. -For all open manipulations of agents that cause splashes or aerosols

HEPA filters required on vacuum lines

Personal Protective Equipments PPE’s: laboratory coats; gloves; face

protection as needed

BSL-1 plus:

Autoclave available

Locking Doors

BSC (as needed)

L Eyewash readily available

Negative pressure in

laboratory (ideal)

Biosafety Cabinet

• What is a Biosafety Cabinet (BSC)?

• Understand Equipment Limitations

• Safe Use of a BSC Why & When to Use a BSC

• SOP for BSC

• BSC Maintenance / Certification Decontamination

What is a Biosafety Cabinet?

• A BSC is a piece of equipment that is only as good as the users understanding of how to appropriately use and maintain it

• There are different Classes of BSC’s Class I – user protection but no product protection Class II – product, user and environment protection

• Provides a ~100ft/min protective air barrier Class III- Air tight - use with Risk Group 3/4 agents

• Fume hoods and laminar flow clean work benches are NOT Biosafety Cabinets.

What is a Biosafety Cabinet

Outline

• What is a Biosafety Cabinet (BSC)?

• Understand Equipment Limitations

• Safe Use of a BSC Why & When to Use a BSC

• SOP for BSC

• BSC Maintenance / Certification Decontamination

Understand Equipment Limitations• Fume Hoods

• Not for use with infectious materials or environmentally dangerous organisms

• Exhaust air not HEPA filtered; not easily decontaminated

• Laminar Clean Air Horizontal

• Air blows through HEPA at rear of work surface across work surface and into face of user

• NOT for use with lab animals, potentially infectious materials, drug formulations.

Vertical• Air blows through HEPA on

top of work surface downward• Air may flow under a sash and

into the room. Some models don’t even have sashes

• Turbulence can distribute aerosols into user’s breathing zone

• Not for use with potentially infectious materials

Reverse-flow• Pull air from front of cabinet

through pre-filter and HEPA at rear

• Used to reduce user’s exposure to animal urine, dander etc. (with PPE)

• Not for work with biohazards (no containment)

Understand Equipment Limitations

• BSC class I Inward airflow

protects worker Exhaust to outside

w/wo HEPA filter No product

protection Not for use with

tissue culture

• BSC class II “Sterile” work

surface Protects worker,

product and environment

For use with RG 2-3 agents

Restrictions for flammables and chemicals depending on type of class II BSC

Understand Equipment Limitations• Class II BSC

Type A2 – 30% exhausted to the room• Not recommended for hard ducting or chemical usage

All Type B cabinets – air flow from the rear grill is discharged into the exhaust system so activities that may generate chemical vapors should be conducted towards the rear of the BSC

Type B3 – 30% exhausted to outside• Minute chemical usage allowed

Type B1 – 70% exhausted to outside• Minute amount of volatile chemicals allowed

Type B2 – 100% exhausted to outside • Small amount of volatile chemicals allowed• Not a good choice for tissue culture work

Outline

• What is a Biosafety Cabinet (BSC)?

• Understand Equipment Limitations

• Safe Use of a BSC Why & When to Use a BSC

• SOP for BSC

• BSC Maintenance / Certification Decontamination

• Resources

Safe Use of a Biosafety Cabinet Must be isolated from other work areas,

lab entry, high traffic areas and away from air ducts

Must be operated properly to minimize risk (more info later in SOP section)• Sash must be kept at manufacturers

recommended height while in use with biological agents.

Safe Use of a Biosafety Cabinet Flames are not a constant temperature

and therefore can cause air turbulence inside a BSC• Compromise protective air barrier integrity

Are not spark/fire proof. Gas Bunsen burners should not be used in BSC’s• 70% recirculated (Class II type A2)

Uncombusted gas (yellow flame) captured in plenums. UV light – microscopic holes in gas tubing. Net result high explosion hazard

Bunsen Burner - BSC Fire

Safe Use of a Biosafety Cabinet UV lights

• Cause skin cancer and cataracts. Therefore eyes and skin should not be exposed.

• Not reliable as primary decontamination Line of site Poor penetration UV light turns on at 80 micro watts /square cm UV light only kills at 160 micro watts/square cm

• Secondary decontamination 15 minutes generally sufficient More than 15 minutes eats up plastics and glue

in HEPA filter and in long term can destroy BSC integrity and cause cabinet leak test to fail

Safe Use of a Biosafety Cabinet

• Do’s Keep supplies to a minimum

and ~6” from sides Discard all infectious materials

inside the BSC (disinfectant) Understand how the BSC

works Turn the cabinet on for

manufacturer recommended time before using it

Inform your supervisor if you are immuno-compromised

Wear a mask if you are coughing or sneezing

• Don’ts Rely on the UV light to

decontaminate Put anything on the front grill Move quickly Use sweeping motions Keep supplies close to the sides

and back Move the sash below or above

the recommended standard Use a gas flame in the hood Remove material before

disinfecting. When working with potentially infectious material

FDS

Safe Use of a Biosafety Cabinet

1. What happens if the front sash is up too high?

Safe Use of a Biosafety Cabinet

1. Question: Front sash up too high

Answer: Down flow of clean (HEPA filtered) air will provide more pressure than inflow of air through the front sash and safety is compromised i.e. agent contamination of the user and environment is likely.

FDS

2. What Happens If The Front Sash Is Down Too Low?

Safe Use of a Biosafety Cabinet

2. Question: Front sash down too low.

Answer: In flow of air through front sash is stronger than down flow of air. Net result is compromised product integrity, i.e. outside air flows into the cabinet providing a source of potential contamination.

FDS

3. What Happens If Objects Are Placed On The Front Grill Of The BSC?

Safe Use of a Biosafety Cabinet

3. Question: Effect of objects placed on the front grill.

Answer: The front grill is one of the dirtiest areas of the entire lab so placing items on this grill is not a good idea. Additionally this interferes with air balance and causes too much down flow of air which creates safety issues, i.e. release of agent outside of the cabinet.

FDS

4. What Happens When Your Arm Moves Over The Work Inside The BSC?

Safe Use of a Biosafety Cabinet4. Question: Arm moves over work area.

Answer: Down flow air moves contaminants on your arm/lab coat down onto the work surface compromising product sterility.

Note: If you are right handed the waste container should be on the right side of the BSC.

FDS

5. What Happens When People Walk Behind The BSC?

Safe Use of a Biosafety Cabinet

5. Question: Person walks behind the Biosafety Cabinet.

Answer: Protective air current is ~100 feet / minute = a little over one mile an hour. People walk at 2-3 miles per hour and this can disrupt the protective air current of the biosafety cabinet.

Outline

• What is a Biosafety Cabinet (BSC)?

• Understand Equipment Limitations

• Safe Use of a BSC When to Use a BSC

• SOP for BSC

• BSC Maintenance / Certification Decontamination

• Resources

When to Use a BSC

• Product Protection Tissue Culture

• User /Environmental Protection Aerosolizing activities with Risk Group 2

agents.

Aerosol Producing Activities

• Sonication, blending, mixing, vortexing

• Almost any liquid manipulation

• Centrifuging

• Pouring

• Pipetting

• Opening containers at non-ambient pressures, (e.g. fermenters, freezer vial)

• Loading syringes and injections

Aerosol Producing Activities (cont.)

• Intranasal inoculation of animals

• Changing animal bedding

• Harvesting tissue, eggs

• Tissue grinder/homogenizers

• Lasers

• Cell sorters

• Necropsy

Aerosol Control Procedures

• Biosafety Cabinet (BSC) properly maintained, used and certified

• No blow-out of pipettes

• Sidewall contact when delivering liquids

• Use of aerosol-minimizing tips

• Secondary enclosure for sonicators, etc.

• Capped centrifuge tubes, sealed cups, carriers and rotors

Outline

• What is a Biosafety Cabinet (BSC)?

• Understand Equipment Limitations

• Safe Use of a BSC Why Use a BSC

• SOP for BSC

• BSC Maintenance / Certification Decontamination

• Resources

Why Use a BSC? – User Protection

Account for 50% of LAI’s (Lab Acquired Infections)

1. Inhalation exposure to infectious aerosols

Account for 20% of LAI’s

1. Parenteral Inoculation (to include animal bites and scratches)

2. Spills, splashes, skin and mucous membrane exposure

3. Ingestion

Outline

• What is a Biosafety Cabinet (BSC)?

• Understand Equipment Limitations

• Safe Use of a BSC Why & When to Use a BSC

• SOP for BSC

• BSC Maintenance / Certification Decontamination

• Resources

BSCStandard Operating Procedure

Start up Procedure

• Pre-plan to minimize supplies and movements in and out of the cabinet

• Turn on for manufacturer’s recommended time prior to use

• Ensure that back grill paper catch is clear

Start up Procedure

• Verify air flow

• Verify drain valve is closed

• Disinfect cabinet surfaces

• Place all materials inside the cabinet (disinfect as appropriate)

• Verify view screen height is appropriate

BSCStandard Operating Procedure

Work in the Cabinet

• Wash hands and arms

• Do NOT cover the front or back grill with anything

• Use slow movements

• Avoid door and people movement beside cabinet

• Avoid open flames

Work in the Cabinet

• Place pipettes exposed to infectious materials in disinfectant inside the cabinet before removing

• Safe work area is 6 inches from sides of cabinet

• Enclose all potentially contaminated material & disinfect surfaces before removing from BSC

• Disinfect work surfaces

Standard Operation Procedure - BSC

• Develop from Manufacturer’s guidelines

• Baker SOP available in BSL-2 Biosafety Manual Template (supplemental resource D) http://www.bio-safety.wsu.edu/forms.asp

Outline

• What is a Biosafety Cabinet (BSC)?

• Understand Equipment Limitations

• Safe Use of a BSC Why & When to Use a BSC

• SOP for BSC

• BSC Certification

• Resources

BSC Certification

Certification

• Must be certified every year / after installation / and after the BSC is moved

• NSF certifier to provide certification to NSF standards requires 2 HP compressor *Pump designed for aerosol

challenge

Certification

• Check air-flows 100-110 ft/min in-flow (A2) 75 ft/min in-flow (A1) 65-75 ft/min down-flow

• Check HEPA filter Particle generator* – 18-20

lbs pressure to produce 0.3 micron particle size (99.9% efficiency test)

Outline

• What is a Biosafety Cabinet (BSC)?

• Understand Equipment Limitations

• Safe Use of a BSC Why & When to Use a BSC

• SOP for BSC

• BSC Maintenance

• Resources

BSC Maintenance

Maintenance

• Keep the cabinet clean

• If cabinet is off keep front sash down to minimize dust

• Clean up spills that get under the cabinet immediately http://www.bio-safety.wsu.edu/biosafety/

• Disinfect before & after use

• Bleach pits stainless steel

• Monitor minihelic gauge

Maintenance

• Don’t store materials in the hood

• Verify that the front grill/back grill and paper catch are unobstructed

• Once a month disinfect the bottom of the hood and then drain

• >10-12 year old BSC’s - stop stocking parts

Outline

• What is a Biosafety Cabinet (BSC)?

• Understand Equipment Limitations

• Safe Use of a BSC Why & When to Use a BSC

• SOP for BSC

• Decontamination

• Resources

Decontamination• Microorganisms can be grouped in terms of

resistance to disinfectants.

Highly Susceptible Susceptible Resistant

Highly Resistant

Extremely Resistant

Microbial Chemical Resistance

Highly Susceptible Mycoplasmas

Susceptible Gram +&- bacteriaFungal Spores

Resistant Non-Enveloped Viruses

Mycobacteria

Highly Resistant Bacterial Endospores

Protozoal Oocytes

Extremely Resistant Prions

Effectiveness of Chemical Disinfectants• Effectiveness is influenced by:

Composition of chemical disinfectants Concentration of microorganisms and

chemical disinfectants Contact time with the Disinfectant Presence of organic matter Presence of interfering substances Temperature at which they are used

Summary of Disinfectant ActivitiesDisinfectant

Disinfection Level

Bacteria Lipophil. Viruses

Hydro-Philic

Viruses

M. tuberculosis

Fungi

Comments

Quaternary Ammonium (0.5-1.5%)

low + + - - +/ Ineffective against bacterial spores. May be ineffective against Pseudomonas and other gram negative bacteria; recommendation limited to environmental sanitation

Alcohols (ethyl and isopropyl) 60-85%

Intermed. + + - +/ + Ethyl or isopropyl alcohol at 70-80% concentration is a good general purpose disinfectant; not effective against bacterial spores. , high concentrations of organic matter diminish effectiveness; flammable

Phenolics(0.4%-5%)

Intermed. + + +/- + + Not sporicidal; phenol penetrates latex gloves; eye/skin irritant; remains active upon contact with organic soil; may leave residue

Chlorine (100-1,000 ppm)

Intermed. + + + +/ + Not generally sporicidal; inactivated by organic matter; fresh solutions of hypochlorite (chlorox) should be prepared weekly; corrosive; irritating to eyes and skin

Iodophors (30-1,000 ppm iodine)

Intermed. + + + +/ +/ Recommended for general use. Wescodyne

diluted 1 to 10 is a popular disinfectant for washing hands. Inactivated by organic matter

Glutaraldehyde (2-5%)

high + + + + + Used to sterilize surgical instruments that can not be autoclaved; strong odor; use with adequate ventilation. Not for use on environmental surfaces. Because it is a sensitizer and causes asthma it is not recommended for laboratory use.

Case Study – Putting it all Together

• Work with M. bovis. rDNA M. bovis culture

• Agent Risk Group http://www.absa.org/riskgroups/bacteriasearch.php?genus=Mycobacterium

• Risk Assessment

• Appropriate BSL

• Special Considerations

Practicing Science Safely Means That Before Any Activity...

YOU KNOW the risks • MSDS

YOU KNOW the worst things that could happen • Sick/Die

YOU KNOW AND USE the prudent practices, protective facilities, and protective equipment needed to mitigate the risks• Risk Assessment

YOU KNOW what to do• Follow Biosafety Manual

A Complete Risk Assessment includes• Agent

Characterization (Risk Group RG)

• Personnel Factors (experience)

• Work Activity Factors

• Environmental Factors

• Equipment Factors

• Risk Consequences

• Probability Profile

A- Agent Characterization• Pathogenicity of material – disease incidence and severity

• Routes of Transmission – parenteral, airborne or ingestion

• Agent Stability – ease of decontamination

• Infectious Dose – LD50

• Concentration – infectious material/volume & working volume

• Origin of material - Wild Type, exotic, primary cells

• Availability of effective prophylaxis – Hepatitis B vaccine

• Medical surveillance – effectiveness & availability of treatment

A- Agent Characterization• Pathogenicity of material – 4th most common LAI, MDR strain

used increases the risk.

• Routes of Transmission – parenteral, airborne & ingestion

• Agent Stability – Resistant to many disinfectants

• Infectious Dose – 10 bacilli by inhalation

• Concentration – cultures grown on plates

• Origin of material - Wild Type

• Availability of effective prophylaxis – INH (Isoniazid) available, ineffective for most MDR strains

• Medical surveillance – treatment of MDR Mycobacterium problematic

B - Personnel Factors• Biosafety training

• SOP training

• Health Assessment (immunosuppressed)

• Experience with the Agent (B, I, A) Beginner, Intermediate, Advanced

• MSDS read

• Experience with the procedures (B,I,A)

• Use of PPE training

• Job hazard analysis undertaken

• Allergies (animal, environmental)

C- Work Activity Factors

• Aerosol generating potential

• Potential for self-inoculation (needle stick, lesion)

• Sample origin and concentration

• Volume of pathogen used• Animal use (types,

potential viral shedding, bites and scratches)

• Replication competency • Recombinants• Cell line characteristics

• Toxin production (y/n, MSDS))• Modification of pathogen (y/n, result

/ implication)• Vector use (y/n, describe) • Inventory Records• Contingency plan (exposure,

accidental releases / spills)• Techniques – cryogenics, cytometry• Disinfectant used as directed

D – Environmental Factors

• Level of containment available• Degree of monitoring of containment factors• Impact of external activities• Biosecurity (access and inventory control)• Availability and status of emergency support • Housekeeping and Trades Personnel• Access by public

E – Equipment Factors

• Equipment Maintenance (frequency, status)• Manual • Reservoirs empty &disinfected• Standard Operating Procedures• Location of use• Ventilation Consideration

F - Rating of Risk Consequences• Rating of Risk Consequences

Class Rating Consequences

4 Catastrophic People: fatalities, evacuation outside site areaEnvironment: irreversible, long-term damage outside site areaBusiness: total loss: > $2 million Interruption: > 2 months image: severely damaged, > 1 week, national

3 Critical People: serious injuries, effects outside site areaEnvironment: reversible, short-term damage outside site areaBusiness: total loss: > $100,000 - $2 million Interruption: > 2 – 8 weeks image: damaged, > 1 week, regional

2 Marginal People: minor injuries, annoyance outside site areaEnvironment: only site area effectedBusiness: total loss: > $5,000 - $100,000 Interruption: > 1 – 2 weeks Image: < 1 week, local

1 Negligible People: no effectsEnvironment: only building effectedBusiness: total loss: < $ 10,000 Interruption: > 1 week Image: no effects

F - Rating of Risk ProbabilityTable 2 - Rating of Probability

Class Rating Probability Definition

A Frequent More than once a year Likely to occur repeatedly in life cycle system

B Moderate Once per year Likely to occur several times in life cycle system

C Occasional Once in 5 years Likely to occur sometime in life cycle system

D Rare Once in 25 years (e.g. once in the life cycle of the system)

Not likely to occur in system life cycle, but possible

E Unlikely Once in 100 years Will occur once in a lifetime of a site

F Very Unlikely Once in 1,000 years Almost impossible to occur

G -Probability ProfileRisk Profile/Probability

Consequences

Probability

Negligible 1

Marginal2

Critical3

Catastrophic4

FrequentA

ModerateB

OccasionalC

RareD

UnlikelyE

Very UnlikelyF

Resources - People

• WSU – Biosafety Pullman and offsite campuses Lorraine McConnell 509-335-4462

lorrmcc@wsu.edu

• WSU- EH&S Pullman Mike Kluzik 509-335-9553

mkluzik@wsu.edu

 

Resources• Biosafety in Microbiological and Biomedical

Laboratories: http://www.cdc.gov/od/ohs/biosfty/bmbl5/bmbl5toc.htm

• American Biosafety Association: http://www.absa.org/XriskgroupsX/index.html

• Canadian Biological MSDS’s: http://www.phac-aspc.gc.ca/msds-ftss/index.html

• WSU Biosafety Web Page: http://www.bio-safety.wsu.edu/biosafety

Web Resources - BSC• CDC / NIH Guidelines on Biosafety Cabinets:

http://www.ors.od.nih.gov/ds/pubs/bsc/contents.html

• AIHA – BSC Safety Information: http://www2.umdnj.edu/eohssweb/aiha/accidents/

BSC.htm#Biosafety%20Cabinet

• CDC / NIH 2nd ed. “Primary Containment for Biohazards”: file:///Z:/Biosafety%20Cabinets/CDC-Primary

%20Containment%20for%20Biohazards%20(BSC's).htm

• Ohasis – Office of Health & Safety Primary Containment for Biohazards Section V. file:///Z:/Biosafety%20Cabinets/Primary%20Containment

%20for%20Biohazards.htm

Web Resources - BSC• CDC – Laboratory Biosafety Guidelines 3rd edition:

http://209.85.173.104/search?q=cache:h373QUWmYfEJ:www.phac-aspc.gc.ca/publicat/lbg-ldmbl-04/pdf/lbg_2004_e.pdf+CDC+2007+3rd+edition+of+Selection+Installation+and+Use+of+BSC&hl=en&ct=clnk&cd=5&gl=us&client=firefox-a

• ABSA Position Paper on the Use of Ultraviolet Lights in Biological Safety Cabinets: http://209.85.173.104/search?

q=cache:tbuGD9FOadAJ:www.ehs.umass.edu/ABSA%2520UV%2520light%2520paper.pdf+position+paper+on+the+use+of+ultraviolet+lights+in+biological+safety+cabinets&hl=en&ct=clnk&cd=1&gl=us&client=firefox-a

Web Resources - BSC

• BMBL Biosafety Cabinet Appendix A (4th ed): http://www.cdc.gov/od/ohs/biosfty/bmbl4/b4aa.htm

• The Baker Company http://www.bakerco.com/resources/intro.php

• NUAIRE How to select a BSC http://www.nuaire.com/products/

biological_safety_cabinets/select_a_bio_cabinet.htm

• NSF Accreditation Standards http://www.nsf.org/business/biosafety_accreditation/

index.asp?program=BiosafetyCabCert

Regulations• OSHA Bloodborne Pathogen Standard:

http://www.osha.gov/SLTC/bloodbornepathogens/index.html

• Centers for Disease Control and Prevention (CDC): http://www.cdc.gov/od/sap/

• NIH Recombinant DNA Guidelines: http://www4.od.nih.gov/oba/rac/guidelines/guidelines.html

• CDC Interstate Shipment of Etiological Agents: http://www.cdc.gov/od/ohs/biosfty/shipregs.htm

• CDC Etiologic Agent Import Permit Program: http://www.cdc.gov/od/eaipp/

• APHIS – USDA: http://www.aphis.usda.gov/vs/ncie/

THANK YOU