Biosafety

BioSafety
Dr Peter Jones,
Biological Safety Officer,
School of Biomedical Sciences

Dr John Sutherland,
University Safety Officer
Outline
Biological Agents
Classification
Lab facilities
Safety Cabinets; Fumigation; Autoclaves; Disinfectants; Waste disposal; Cryogens, Latex
Transport
Genetic Modification
Cell Culture
Training
Legislation
Control of Substances Hazardous to Health Regulations 1999 (COSHH)
ACOP - Biological Agents
ACDP Guidance
4th Edition - categorisation/containment
blood-borne viruses
Transmissible Spongiform Encephalopathies (TSE)
Genetically Modified Organisms (Contained Use) Regulations 2000
HSE Compendium of Guidance
Biological Agents
COSHH Definition
Any of the the following if they can cause infection, allergy, toxicity or other human harm.
micro-organism
cell culture
human endoparasite
COSHH Requirement
Assess health risks from the BA
worker, co-workers,
cleaners, maintenance, waste handlers.
Prevent or control exposure
Maintain control measures
Safety cabinets
PPE - lab coats
Classification
ACDP 4th Edition 1995
“Categorisation of Biological Agents According to Hazard and Categories of Containment.”
HSE Approved List - schedule to COSHH
Bacteria, Viruses, Parasites & Fungi
Hazard grouping 1 - 4 (low to high)
Containment facilities/lab standards (1 - 4)
List identifies lab facility for the organism
Biological Agents
Hazard Group 1
Unlikely to cause human disease
Hazard Group 2
Can cause disease
May be a hazard to employees
Unlikely to spread to community
Prevention or treatment available
Bacillus cereus,
Clostridium spp,
Most E. coli,
Pseudomonas aeruginosa,
Proteus vulgaris, Staph aureus.
Hazard Group 3
Can cause severe human disease
Serious risk to employees
May spread to community
Prevention or treatment available
Anthrax; Brucella abortus/canis/suis; E.coli O157 Mycobacterium bovis/leprae/tuberculosis; Salmonella typhi/paratyphi; Yersinia pestis.
HIV; SIV; Hepatitis; Hantaan
Plasmodium faliciparum, Trypanosoma cruzi
Hazard Group 4
Virusus such as
Lassa fever
Congo heamorrahagic fever
Ebola
Marburg
Variola

CANNOT BE USED HERE
Containment Facilities - level 2
Bench - impervious, washable, chemical resistant.
Floor - coved, continuous, sealed.
Safety cabinet for infectious aerosols.
Restricted access, door kept closed.
Safe, secure storage of organisms.
Clean lab coats - side/back fastening.
Disinfection and waste disposal procedures.
Waste to autoclave in robust spill/leak proof containers.
Wash-hand basin by the door.
Containment Facilities - level 3
Self-contained unit, separate from other activities - own autoclave, incubators etc.
Entrance lobby and change area.
Designated lab coats - separate from other lab coats.
Sealable for fumigation.
Negative pressure to corridor - mechanical ventilation.
Training in use of facility
Microbiological Safety Cabinets
Class 1 to 3
Classes DO NOT relate to containment level!!
Class 3 - highest protection
Class 1 - good general protection
Class 2 - can be OK - combines protection of work and worker against contamination.
Class 1 Cabinet
Open fronted design protects worker by continuously drawing air in through front.
Simple, cheap, effective.
Suitable for Hazard Group 2 work.
Also acceptable for lower risk Hazard Group 3 work.
Class 2 Cabinet
Open fronted protects worker and the work from external contamination.
Air drawn down and filtered before directing into work area as a clean vertical laminar flow.
Need good working practice.
Air flow can be upset by external air movements.
Class 3 Cabinet
Totally enclosed cabinet with operations conducted through gloves attached to glove ports.
HEPA filtered air drawn from rear and extracted.
Very high protection but restricted dexterity.
Suitable for high risk Hazard Group 3 - eg highly concentrated samples
Laminar Flow Hoods
DO NOT confuse laminar flow hoods with Microbiological Safety Cabinets.
LFHs draw clean filtered air vertically or horizontally across the work to protect the work from external contamination.
There is no worker protection as there is no inward air flow - horizontal units direct air towards the operator!!!
Use of Safety Cabinets
Run for a while before use to stabilise airflow.
Check airflow indicator to confirm OK.
Set up cabinet with all equipment needed.
Do not overcrowd cabinet.
Work in centre of open fronted cabinets as far as is possible.
Do not work closer than 15cm to front.
Keep clean and dirty materials separate.
Disinfectants available. Discard jar inside cabinet for HG3. Clean and disinfect surfaces at end.
Safety Cabinet Maintenance
Performance checked on
installation
annually - level 2 lab
6-monthly - level 3 lab
Test sticker on front.
Inspection certificate issued.
Filters need occasional replacement.
Fumigate cabinet and filters before-hand.
Disinfectants - selection
Type
Spectrum of activity
Specific activity for different micro-organisms
Circumstances
Dirty or clean - organic load
intra or extracellular viruses
Chemical incompatability
Temperature, pH, hardness of water.
Disinfectants - selection
Surfaces to be disinfected
Acids, alkalis, hypochlorites, electrolytes can corrode stainless steel and other metals.
Organic solvents can damage plastics.
Powder form, or gelling, to sprinkle on spills, or soak a tissue in contact with the spill.
Hazardous properties
Can be toxic or corrosive.
Gluteraldehyde is a powerful sensitiser.
Reaction products, eg Chlorine.
Disinfectants - selection
Disinfectants - selection
Disinfectants - selection
Phenolics - hycolin; stericol; clearsol
Protein denaturant - lysis of micro-organism
Choice for Mycobacterium.
Damages plastics.
Disinfectants - selection
Chlorine-based hypochlorite;chloros;presept
Rapid action - Protein denaturation
Chlorine produced if mixed with acid
Carcinogens if mixed with formaldehyde
General use 1,000-2,500 ppm
Discard containers 5,000-10,000 ppm
Spills 20,000 ppm
More rapid decomposition at lower dilution.
Disinfectants - selection
Alcohols - 70% ethanol; 60% propanol
Relatively poor efficacy.
Susceptible to interference.
Use on physically clean surfaces.
Flammability risk.

Disinfectants - selection
Aldehydes - cidex; Gluteraldehyde; Formaldehyde
Irritant and toxic
Unsafe for general use-some specific applications
gaseous fumigations
special sensitive equipment under contained conditions
Disinfectants - selection
Surface active agents - cetrimide; tego
relatively safe
inactivated by organic material and anionic detergents eg soap.
Disinfectants - selection
Peroxygen compounds - Virkon
Broad spectrum of effectiveness
1% working solutions are relatively safe
Colour indication
Stable for 7 days
Can attack steel surfaces
Use of Disinfectants
Stability of working solutions.
Effectiveness decays with time.
Some (eg virkon) have colour indicator.
Label with expiry date.
Contact time.
Check concentration and contact time necessary for your conditions - suppliers data.
Use of Disinfectants
Validation.
With HG3 organisms, or working off-spec with HG2 organisms, validate the effectiveness of the disinfection protocol experimentally.
Formalise into lab disinfection procedures.
Fumigation
MSCs prior to servicing etc
CL3 labs after major spillage - smoke test first.
Formaldehyde - toxic; needs humid atmosphere.
Remove chlorine-containing compounds from area - lung carcinogens produced.
Written protocol
Warnings displayed
Notification might be needed to Engineers in Med School.
Fumigant exhausted without exposure.
Autoclaves
Written operating procedure
Training and authorisation needed
Visual check of seals and steam leaks pre-use
Protective clothing - lab coat; impervious apron; heavy duty gauntlets; face visor, robust shoes
Clear separation of clean and dirty items.
Ensure steam penetration throughout the load
Ensure materials are suitable
NO RADIOACTIVE MATERIALS
Autoclaves
Biohazard (Clinical) Waste
Requirements
Identification
Segregation
Packaging
Yellow bags
Label source - dept name tape
3/4 full max.
Seal - tie, knot, proprietary clip.
Remove to secure collection point
Final Disposal - Incineration
Infective wastes - autoclave first
Centrifuges
Use sealed buckets or rotors
Check seals before use
CL2 & 3 - open in safety cabinet
Clean and disinfect centrifuge and rotor after use.
Some disinfectants attack metal rotors!
Transport of Biological Material
Regulations for road, rail and air.
Post Office Requirements.
Classification, labelling and packaging by competent person.
Hazard Group 2, 3 and 4 pathogens/GMOs.
Package - UN approved to prevent release
Special package if dry-ice used.
Shipper’s declaration for air transport.
advise use shipping agent.
Transport of Biological Material
Mini Tube
Biofreeze - 1ltr
Unscreened human tissue and fluids
Risk of “hidden” pathogens.
CL2 if unknown, CL3 if known HG3 present.
Blood-Borne Viruses are HG3 (usually)
Very low aerosol risk - cuts, scratches, injection
Vaccination available for HBV - contact Occ Health.
Contamination of skin, mucous membranes, work area.
Designate area, written protocols followed,
Avoid sharps and glassware,
Cover cuts with waterproof plasters, wear gloves,
Rigorous decontamination procedures,
MSC if aerosols produced - mixing, shaking, sonication
Prions/TSE agents
Hazard group 3 - human TSEs
Work at CL2 PLUS
dedicated room
eye protection
gloves, gown and apron
dedicated equipment
MSC for aerosol-producing processes
Special decontamination
- autoclave 134C/18min; 20,000ppm chlorine for 1hour
- final disposal by incineration
Report accidents involving introduction through the skin.
Latex Allergy
latex can cause skin/respiratory sensitiation.
1% of population may react.
history of asthma, dermatitis, eczema, hayfever
Immediate
local or generalised spots and swelling (5-30 mins)
Delayed (peaks 24-48 hours then subsides)
red rash on back of hands / between the fingers
skin may become leathery and develop blisters
Seek advice from Occ Health if skin/resp symptoms
Cryogenic Liquids
Asphyxiation
<18% O2 (spills > 143ml LN/m3 : 1:700)
lifts, confined spaces,
Cold Burns
eye protection - goggles/visor/s.specs
hands - non absorbent insulated gloves eg leather. Sleeves over ends.
Transport
separate from driver, secure, labelling
END
Hazardous Substances - GMO`s
Genetically Modified Organisms (Contained Use ) Regulations 2000
Classification into Hazard Group GM Risk Assessment
Risk factor
Containment level
GM Safety Committee
Biological Safety Officer
Health Surveillance (via Occ. Health)