1. Basic Biosafety Principles
EH&S Academy
Brenda J. Wong, UCSD Biosafety Officer
October 2009

2. What is Biosafety?
Safety from exposure to Infectious Agents
Smallpox

3. Did the Plague Kill Illinois Scientist?
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.

4. 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"

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

6. 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

7. Biosafety Issues Laboratory Safety Bloodborne pathogens (BBP)
Recombinant DNA (rDNA)
Biological waste disposal
Infectious substance and
diagnostic specimen shipping

8. 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

9. Biohazardous Materials
Viruses
Bacteria
Fungi
Chlamydiae/Rickettsiae
Prions
Recombinant DNA

10. Biohazardous materials
Transgenic Plants, Animals and Insects
     
   

11. Transgenic Insects

12. Biohazardous Materials
Human and Primate Cells, Tissues, and Body Fluids
Brain Tissue from Demented Patients
Viral Vectors
Replication deficient viruses

13. Biosafety Concepts 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.
Lab. Animal Facilities
The 4th edition of the BMBL (1999) contained the first government writings about security concepts for Select Agents. By inclusion, these ideas can be used by all laboratories that store and use pathogens, particularly those that might be perceived as or used for terrorist purposes.
HHS Publication No. (CDC)

14. The New BMBL
Early print edition….
Emphasis on “Risk
& Containment”

15. Biosafety Concepts The BMBL
The BMBL continues
to be published by the
CDC and the NIH
5th edition is now at the printers

16. Are the NIH Guidelines Optional?
“Guidelines” does not mean “optional”
They are a term and condition of NIH funding for recombinant DNA research.
Now although they are called ‘Guidelines’ we don’t want to give you the impression that they are optional. You can’t pick and choose which sections to follow based on what sounds like a good idea or seems to much trouble to deal with.
The Guidelines are a term and condition of NIH funding for recombinant DNA research and failure to comply with them can affect the funding for recombinant DNA work at an institution.
From Kathryn Harris, NIH, OBA

17. 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

18. 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 (ABSL)
Laboratory animal facilities
Animal models that support research
Guidelines for working safely in animal research facilities

19. 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

20. 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

21. 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.

23. Laboratory Design
“Warehouse Type Lab”

24. Discussion
What are some of the negatives and positives of this open lab concept?

25. 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

26. Risk Group 1 Agents E.coli K-12 Transgenic Plants Plasmids Fungi Mold
Yeast

27. 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

28. Risk Group 2 Agents Human or Primate Cells Herpes Simplex Virus
Replication Incompetent Attenuated Human Immunodeficiency Virus
Patient specimens

29. 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

30. 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

31. 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

32. Risk Group 3 Agents Human Immunodeficiency Virus
Mycobacterium tuberculosis
Coxiella burnetii

33. 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

34. 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

35. 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”

36. 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

37. UCSD’s BSL-3

38. Biosafety Level-4 Working in High Containment
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

39. Biosafety Level 4 Lassa Fever Virus Ebola Hemmorrhagic Fever Virus
Marburg Virus
Herpes B Virus

40. 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

41. Animal Biosafety Level-4 Working in High Containment

42. 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

43. 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)

44. Animal Containment Points
CDC
CDC & UCSD
Courtesy of Paul Vinson, CDC

45. 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?

47. Risk Assessment
In-Vitro
Human Clinical
Trial
In-Vivo

48. 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?

49. Risk Assessment, con’t
Tom Pugh
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?

50. 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?

51. 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?

52. 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?

53. First Aid Measures Splash to Eye or Needlestick Injury
Rinse thoroughly for 15 minutes at the eyewash or sink
Call Occupational Medicine 
Call EH&S to report exposure –

54. What Helps?

55. Resources
UCSD Biosafety:
Biological Safety MSDS:
NIH BMBL:
See Handout for additional resources..

56. Questions? Brenda Wong Bwong@ucsd.edu 858 534-6059 Cell: 858 583-3277
Additional Resources:
Biosafety Resources.doc