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An Introduction to General Laboratory Safety & Biological safety

How Safe do you think a Laboratory is? Is it a Danger Zone?

Mike Mackett PhD, DipOSH, CMIOSH University of Hong Kong Biological Safety Officer

Hazard

“The potential to cause harm”

Risk

“The likelihood (probability) of harm actually occurring and the severity of its consequences”

A Few Definitions

Safety is basically concerned with controlling the risks

Hazard identification: What are the major hazards in a laboratory?

Slips, Trips, Falls Electric Shock

Heat/Cold -Autoclaves, Freezers, Liq N2

Pressure (positive and negative) – Gas cylinders Musculo-skeletal - Manual Handling, computer use etc

Glassware, sharps/ knives, needles Chemicals – Fire, Burns, Toxicity, Carcinogenicity

Biological agents - Infection Radiation, Lasers

Noise e.g. From machinery, Biohazard suits etc.

Basic Laboratory Safety Rules • Observe the warning signs and symbols. • Learn about the Dos and Don’ts. • Assess risks before starting work. • Know the location of and how to use the emergency

equipment. • Know the types of personal protective equipment

available and use them e.g. labcoats, gloves, goggles and respirators.

• Always be alert to unsafe conditions and actions and call them to the attention of your supervisor. Safety is the responsibility of everyone - don’t expect someone else to sort it out.

• Use equipment only for its designed purpose

A Selection of Safety Related Signs

Printable public domain signs available at:- http://chemistry.about.com/od/healthsafety/ig/Laboratory-Safety-Signs/

Mandatory-Action

Warning

Prohibition

First Aid Box

Know the location of and how to use the emergency equipment (who is the first aider/firewarden in your lab?)

Fire Extinguishers Hosereels/Exits

Eyewash Station /Shower

Chemical spill kits

Basic Laboratory Safety Rules • Observe the warning signs and symbols. • Learn about the Dos and Don’ts. • Assess risks before starting work. • Know the location of and how to use the emergency

equipment. • Know the types of personal protective equipment

available and use them e.g. labcoats, gloves, goggles and respirators.

• Always be alert to unsafe conditions and actions and call them to the attention of your supervisor. Safety is the responsibility of everyone - don’t expect someone else to sort it out.

• Use equipment only for its designed purpose

Biological Safety – Biosafety

The aim of Biosafety is to reduce or eliminate accidental exposure to, or release of, infectious agents (including Bacteria, Fungi, Viruses, Parasites and cell cultures)

What are the major biological hazards in laboratories?

Risk of Infection from:- • Deliberate culture of various agents • Virus vector work – e.g. AAV, Adenoviruses, Retroviruses • Clinical Samples, bloods, histology specimens, FACS etc • Zoonosis (agents that can infect animals and humans) • Cell culture – risk mainly from adventitious agents that

may be present in primary cultures • Clinical waste

(risks may be altered by Genetic Modification)

Biological experiments with Carcinogenic, Teratogenic or Toxic chemicals/agents

Splashes and sprays 188 Needlesticks 177 Sharp objects 112 Animal or ectoparasite bite/scratch 95 Mouth pipetting 92 Other, unknown 39 Total 703 Adapted from Sewell (1995), Clin. Micro. Rev. 8(3) 389–405

Types of accidents associated with laboratory-acquired infections

80% of LAI’s can’t be linked with a known accident – aerosols?

Aerosols Produced from Laboratory Operations

1010 bacteria/ml culture – 10 min Dimmick et al., 1973

Blender, opened immediately on stopping 106

Sonicator, with bubbling 106 Pipetting, vigorous 106 Dropping culture 3 x 105 Splash on centrifuge rotor 105 Blender, opened at 1 minute 2 x 104 Pipetting, carefully 104

Other aerosol generating operations Unscrewing bottle cap Withdrawing needle from septum Vortex mixing Blowing out pipette Animal cage cleaning etc

Risk Group 1 (no or low individual and community risk) Risk Group 2 (moderate individual risk, low community risk) Risk Group 3 (high individual risk, low community risk) Risk Group 4 (high individual and community risk)

WHO Classification of infective microorganisms by risk group

Risk group allocation

Based on degree of hazard to the individual, community and environment.

Degree of Hazard determined by:-

Infectivity Disease or agent Dose Route of Infection Measles 0.2 Inhalation Shigella flexneri 180 Ingestion Vibrio cholerae 108 Ingestion Ease of Transmissibility Result of infection Host Range of agent Availability of treatment/vaccine

Risk Group/ Hazard Group 1 E.Coli K12 Saccharomyces cerevisae Adeno-associated virus Bacillus subtilus Baculovirus

Secondary Schools, Undergraduate teaching etc, Basic research

Risk Group/ Hazard Group 2

8 human herpesviruses - HSV,VSV, EBV, CMV, HHV6,7 & KSHV ETEC, UroTEC Adenovirus (all 50+ human serotypes) Cornybacteria diphtheria Neisseria meningitidis Stapylococcus aureus (including MRSA) Ringworm

Most research laboratories, primary healthcare labs, teaching

Risk Group 3 Vibrio cholera (783 cases and 8 deaths Senegal 05/05) Herpes B virus (upgraded to level 4 in some lists) HCV, HBV, HIV (cf EU/UK and US) E coli O 157 (cf EU/UK and US) Shigella flexnerii Mycobacterium tuberculosis SARS

Some clinical path labs; a specialist laboratory facility

Risk Group 4. Hemorrhagic fever viruses such as:- Rift Valley Fever (264 cases and 109 deaths in Tanzania 05/07); Marburg (423 cases and 357 deaths in Angola 06/05); Ebola (69 cases and 31 deaths in Republic of Congo 09/12); Yellow Fever (849 cases and 171 deaths, Dafur 10-12/12); Smallpox

High spec, specially built, relatively few worldwide but numbers increasing

Controlling the risks

Biosafety Levels 1-4 (Containment Levels)

- Provide increasing levels of personnel and environmental protection; - Guidelines for working safely in laboratories using biological agents and describe combinations of:-

Laboratory Practices and Techniques Safety Equipment including PPE (Primary Barriers) Laboratory Facilities (Secondary Barriers)

Work with a particular agent is assigned to one of these 4 categories

Biosafety Levels 1-4

Laboratory Practices and Techniques Knowledgeable/Responsible supervisor

Personnel Aware of potential hazards and proficient in practices/techniques – training arrangements

Biosafety manual (local code of practice) specific to lab

1. Introduction 2. General procedures 3. Operation of unit 4. Local rules 5. Waste – cradle to grave 6. Staff health 7. Testing and maintenance 8. Emergency procedures

Biosafety Levels 1-4

Laboratory Practices and Techniques

Basic Protective Procedures

Include:- 1. tying long hair back, wearing PPE appropriately (eye

protection, laboratory coats, gloves, closed shoes)

2. always using appropriate pipetting devices (no mouth pipetting)

3. never eating or drinking in the laboratory

4. never applying cosmetics, handling contact lenses or placing objects (fingers-nail biting, pencils etc) in the mouth or touching the face, avoid using mobile phones

Controlling the risks - Safety Equipment

Class II Biosafety Cabinets

Facility Design

Easily cleaned and decontaminated, including benchtops Sink for handwashing Sturdy furniture

Containment levels

Containment Measure 2 3 4

Workplace separation No Yes Yes

Air In and Out HEPA filtered No On output Input and double on output

Access restricted Yes Yes Yes, via airlock key procedure

Sealable to permit disinfection No Yes Yes

Special disinfection procedure Yes Yes Yes

Air –ve to wrt outside No Yes Yes

Efficient vector control Yes (for animal work) Yes (for animal work) Yes

Surfaces impervious to water easy to clean

Yes for bench Yes bench, floor, and walls

Yes bench, floor, walls and ceiling

Illustrative Requirements of Containment Levels

Measures 1-8 of 14 in UK COSSH legislation (2002)

Table 3. Relation of risk groups to biosafety levels, practices and equipment RISK GROUP

BIOSAFETY LEVEL LABORATORY TYPE LABORATORY PRACTICES

SAFETY EQUIPMENT

1 Basic Biosafety Level 1 (BSL1)

Basic teaching, research

GMT None; open bench work

2 Basic Biosafety Level 2 (BSL2)

Primary health services; diagnostic services, research

GMT plus protective clothing, biohazard sign

Open bench plus BSC for potential aerosols

3 Containment Biosafety Level 3 (BSL3)

Special diagnostic services, research

As Level 2 plus special clothing, controlled access, directional airflow

BSC and/or other primary devices for all activities

4 Maximum containment Biosafety Level 4 (BSL4)

Dangerous pathogen units

As Level 3 plus airlock entry, shower exit, special waste disposal

Class III BSC, or positive pressure suits in conjunction with Class II BSCs, double ended autoclave (through the wall), filtered air

BIOSAFETY SUMMARY

The risk group of an organism gives a good indication of which level of containment and what procedures are necessary i.e. A risk group 2 agent requires Biosafety level 2 – e.g. Adenoviruses, Lentiviruses etc

However in many countries they are not always absolutely equivalent e.g. Dengue virus

Other features of the activity may modify the requirements e.g. HIV, HBV simple diagnostics vs growth and purification of high titre stocks

For GM work and activities at class 2 or above a full RISK ASSESSMENT is required by HKU.

Un-labeled containers

Broken glass left on the floor.

Heavy items stored above eye level, including chemicals, equipment and office materials.

Generally, it is prudent to avoid working in a laboratory alone. If you need to work alone, let somebody know so that they can take notice of your situation regularly.

Unattended Operations Plan for interruptions in utility services such as electricity, water, and inert gas. Operations should be designed to be safe, and plans should be made to avoid hazards in case of failure, e.g. the failure of cooling water supply.

Working Alone

Lack of routine maintenance

Exploding cryovials

Needlestick accidents University guidance at :-http://www0.hku.hk/safety/pdf/NSP.pdf Particularly common when handling animals Adopt a stringent sharps policy i.e. •not manually re-sheathing, bending or breaking needles prior to disposal, •needles must not be passed by hand between staff, must be disposed of at the site of use, directly to a sharps bin without removal from any syringe barrel….and •the sharps disposal container must be no more than ¾ full

Poor Housekeeping

Thank You!