Basic Biosafety Principles

EH&S Academy

Brenda J. Wong, UCSD Biosafety

Officer

October 2009

What is Biosafety?

� Safety from exposure to

Infectious Agents

Smallpox

What is Biosafety?

Sunday, Sep. 20, 2009

Did the Plague Kill Illinois Scientist? By AP

(AP / CHICAGO) — The University of Chicago Medical Center says the infection that killed a scientist

may be connected to bacteria he researched that causes the plague.

The university said Saturday that its researcher studied the genetics of harmful bacteria including

Yersinia pestis, which causes the illness. He died Sept. 13. His name and age haven't been

released

The medical center says the bacteria he worked with was a weakened strain that isn't known to cause

illness in healthy adults. The strain was approved by the Centers for Disease Control and

Prevention for laboratory studies.

An autopsy found no obvious cause of death but did find the presence of the bacteria. More tests are

planned. No other illnesses have been reported.

Biosafety in Various Disciplines

Biosafety is related to several fields

� ECOLOGY: referring to imported life forms not indigenous to the region (Reggie the alligator)

� AGRICULTURE: reducing the risk of alien viral or transgenic genes, or prions such as BSE/"MadCow“; reducing the risk of food bacterial contamination

� MEDICINE: referring to organs or tissues from biological origin, or genetic therapy products, virus; levels of lab containment protocols BSL-1, 2, 3, 4 in rising order of danger

� CHEMISTRY: i.e., nitrates in water, PCB levels affecting fertility

� EXOBIOLOGY: i.e., NASA's policy for containing alien microbes that may exist on space samples - sometimes called "biosafety level 5"

Biosafety in Academic Research � Research Universities:

Promoting safe laboratory practices, and procedures; proper use of containment equipment and facilities; provides advice on laboratory design and risk assesment of experiments involving infectious agents, rDNA in-vitro and in-vivo.

Bottom Line: Risk & Containment

Biohazard Symbol

� Charles Baldwin at National Cancer Institute at NIH.

� Symbol to be “memorable but meaningless” so it could be learned.

� Blaze orange – most visible under harsh conditions

Biosafety Issues

� Laboratory Safety

� Bloodborne pathogens (BBP)

� Recombinant DNA (rDNA)

� Biological waste disposal

� Infectious substance and

diagnostic specimen shipping

Biosafety Issues (con’t.)

� Respiratory Protection

� Bioterrorism and Select agents

� Mold and indoor air quality

� Occupational safety and health in the use of

research animals

� Biohazards used in animal models

Biohazardous Materials

� Viruses

� Bacteria

� Fungi

� Chlamydiae/Rickettsiae

� Prions

� Recombinant DNA

Biohazardous materials

� Transgenic Plants, Animals and Insects

Transgenic Insects

Biohazardous Materials

� Human and Primate Cells, Tissues, and

Body Fluids

� Brain Tissue from Demented Patients

� Viral Vectors

� Replication deficient viruses

Biosafety In Microbiological

and Biomedical Laboratories

�“BMBL” (acronym)

�CDC/NIH Publication

�Safety “Guidelines”

�Regulations of Institution receives

NIH funding

�Code of Practice and “Gold”

Standard in Industry anl

�Gold Standard

�Clinical & Research Lab.

Biosafety Concepts

HHS Publication No. (CDC) 93-8395

The New BMBL

� Early print

edition….

� Emphasis on “Risk

& Containment”

The BMBL continues

to be published by

the

CDC and the NIH

5th edition is now at

the printers

http://www.cdc.gov/od/ohs/bio

sfty/bmbl5/bmbl5toc.htm

Biosafety Concepts

The BMBL

Are the NIH Guidelines Optional?

� “Guidelines” does not mean

“optional”

� They are a term and condition of

NIH funding for recombinant DNA

research.

From Kathryn Harris, NIH, OBA

Biosafety Concepts from

the BMBL

Principles of Biosafety

� Practice and Procedures

� Standard Practices

� Special Practices & Considerations

� Safety Equipment

� Facility Design and Construction

� Increasing levels of protection

Principles of Biosafety

Biosafety Levels 1-4 (BSL) � Increasing levels of employee and environmental protection

� Guidelines for working safely in research & medical laboratory facilities

Animal Biosafety Levels 1- 4 (ABSL)

� Laboratory animal facilities

� Animal models that support research

� Guidelines for working safely in animal research facilities

Biosafety Concepts The BMBL

(1) Standard Microbiological Practices � Most important concept / Strict adherence

� Aware of potential hazard

� Trained & proficient in techniques

� Supervisors responsible for:

� Appropriate Laboratory facilities

� Personnel & Training

� Special practices & precautions

� Occupational Health Programs

Biosafety Issues The BMBL

(2) Safety Equipment � Primary Containment Barrier

� Minimize exposure to hazard

� Prevent contact / Contain aerosols

� Engineering controls/ equipment

� Personal Protective Equipment (PPE)

� Gloves, gowns, Respirator, Face shield, Booties

� Biological Safety Cabinets

� Covered or ventilated animal cage systems

Biosafety Concepts The BMBL

(3) Facility Design and Construction

� Secondary Barrier/ Engineering controls

� Contributes to worker protection

� Protects outside the laboratory � Environment & Neighborhood

� Ex. Building & Lab design, Ventilation, Autoclaves, Cage wash facilities, etc.

Laboratory Design

“Warehouse Type Lab”

Discussion

� What are some of the

negatives and

positives of this open

lab concept?

Biosafety Level-1

Concepts of Biosafety Biosafety Level-1 (BSL-1 or ABSL-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

Risk Group 1 Agents

� E.coli K-12

� Transgenic Plants

� Plasmids

� Fungi

� Mold

� Yeast

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

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)

� Biohaz. 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

Risk Group 3 Agents

� Human

Immunodeficiency

Virus

� Mycobacterium

tuberculosis

� Coxiella burnetii

Biosafety Level 3 Working in High Containment

Biosafety Level-3 (BSL-3 or ABSL-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 handwashing

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

� Biohaz. 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

UCSD’s BSL-3

Biosafety Level-4 Working in High Containment

Biosafety Level-4

� Builds on BSL-3/ ABSL-3 practices

� Maximum containment facilities

� Pressurized Containment Suite

� BSL-3 + Class III Biosafety Cabinet

� Chemical decontamination showers

� Liquid effluent collection / decontamination

� No BSL-4 labs exist at UCSD

Biosafety Level 4

� Lassa Fever Virus

� Ebola Hemmorrhagic

Fever Virus

� Marburg Virus

� Herpes B Virus

Biosafety Concepts Working in High Containment

Biosafety Level-4 (BSL-4 or ABSL-4)

� Dangerous/exotic agents

� Life threatening disease

� Aerosol transmission

� Agents of unknown risk

of transmission or health affects

� No known treatment

Animal Biosafety Level-4 Working in High Containment

General Good Lab Technique

� Hygienic Practices

� No Smoking, Eating, Applying cosmetics, lip

balm, contacts

� Wash hands after procedures

� Decontaminate lab bench before and after

work

General Operational Practices

� Proper attire

� Minimum – lab coat, safety glasses, gloves

� Plan your work

� Know in advance what you are working with

� Read available resources (MSDS)

http://www.hc-sc.gc.ca/pphb-dgspsp/msds-

ftss/index.html

Animal Containment Points

CDC - 1957 CDC & UCSD - 2005

Courtesy of Paul Vinson, CDC

Discussion # 2

� Based on what you know about Biosafety

Levels, Practices and Operational Controls,

what are some discussion issues for

conducting Biohazard risk assessments?

� How do you approach risks when

addressing a particular organism?

Risk Assessment

In-Vitro In-Vivo Human Clinical

Trial

Addressing Risk Assessments

� What is the organism?

� Is it Wild-type, attenuated, irradiated, or chemically treated? Look at kill data or kill curves.

� What is the max. concentration, volume, infectious dose?

� What is the work space like?

� Aerosolizing procedures? How do they contain their aerosols?

Risk Assessment, con’t � Are personnel trained? Do

personnel understand the organism, infectious dose and symptoms?

� What are their experimental procedures?

� Will they be transporting the material? Shipping intra, inter-state or international?

� Are they doing tissue culture?

� Do they have adequate containment equipment?

Tom Pugh

Risk Assessment, Con’t

� Are they doing this work in-vivo? Have you consulted and discussed this with the Vets and IACUC to determine special needs and housing?

� Waste issues addressed?

� Pregnancy issues with the organisms?

Risk Assessment, con’t

� Do they share their

Tissue Culture room?

� Do they have more than

1 Biosafety Cabinet?

� Occupational Health

informed and set up to

receive patient or offer

counseling?

Accidental Spills

� Evacuate area, alert personnel and

cordon off so that aerosols may settle

� Don PPE; Cover with paper towels and

apply bleach (1 part bleach : 9 parts water

� Allow 15 – 20 min contact time

� Wipe up working towards center

� Use tongs if broken glass is involved

�Is Recombinant DNA involved?

First Aid Measures

� Splash to Eye or Needlestick Injury

� Rinse thoroughly for 15 minutes at the eyewash or

sink

� Call Occupational Medicine � 619 471-9210

� Call EH&S to report exposure – 858 534-5366

What Helps?

Resources

� UCSD Biosafety: http://blink.ucsd.edu/safety/research-

lab/biosafety/

� Biological Safety MSDS: http://www.phac-

aspc.gc.ca/msds-ftss/index-eng.php

� NIH BMBL:

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

� See Handout for additional resources..

Questions?

Brenda Wong

Bwong@ucsd.edu

858 534-6059

Cell: 858 583-3277

Additional Resources:

Biosafety Resources.doc