Biological Safety in Laboratories the CHALLENGE

Biological Safety in laboratories THE CHALLENGE

Safe Labs

• What is ‘safe’? • Who decides? • How to inform occupants? • How to manage change? Involves &relieson SAFETY LAB/RESEARCH COMMUNITY Hygiene Officer • Chemical Laboratories • Teaching Maintenance, Industrial Hygienists etc. • Fire Prevention, BuildingDesign & Fire • Laboratory Research • Research Field • Radiation & Bio Radiation & Safety Officers

•3 EVERYONE

is responsible for quality and safety SAFETY, HEALTH & ENVIRONMENTAL GOALS ZERO… • Workplace injuries • Workplace illnesses • Related injury/illness at home • Property loss • Unintended environmental damage MANAGEMENT OF HEALTH, SAFETY AND THE ENVIRONMENT . Achieving excellence in providing a healthy and safe working environment . Supporting environmentally sound practices in the conduct of University activities . Comply with all applicable health, safety, and environmental protection laws, regulations and requirements BIOLOGICAL RESEARCH IS ANY EXPERIMENTAL ACTIVITY INVOLVING THE FOLLOWING BIOLOGICAL AGENTS: • Human source materials (includes primary and immortal cell lines, clinical specimens, samples, tissues, etc) • Animals and animal source materials • Plants and plant source materials • Recombinant and synthetic DNA/RNA (including all viral vectors, RNAi technologies, etc). • Microorganisms (bacteria, viruses, fungi, prions, yeast, algae, rickettsiae, parasites, etc) • Biological toxins, allergens The CDC Biosafety in Microbiological and Biomedical Laboratories (BMBL) classifies risk group agents to:

RG1 Agent not associated with disease in healthy adult humans e.g. Bacillus subtilis

RG2 Associated with human disease which is rarely serious and preventive or therapeutic interventions are often available e.g.Staphylococcus aureus, influenza viruses type A, B, and C

RG3 Serious or lethal human disease; preventive or therapeutic interventions may be available e.g. Mycobacterium tuberculosis

•RG4 Serious or lethal human disease; preventive or therapeutic interventions are usually not available e.g. Ebola Quick question Which risk group do you think the avian flu virus would belong to?

Resources for assigning risk group/biosafety level 1. The NIH Guidelines Appendix B assigns risk groups to some biological agents. 2. The American Biological Safety Association (ABSA) website provides a searchable database of biological agents and their assigned biosafety levels by country. BIOSAFETY LEVELS BL-1: Agents are not known to cause disease BL-2: Agents are associated with human disease BL-3: Agents are associated with human disease and are potentially transmitted as aerosols BL-4: Agents of life threatening nature BIOSAFETY LEVEL 1 (BL-1)

Use BL-1 when working with: • Well characterized agents • Agents that are not known to cause disease in healthy humans • Agents that are of minimal hazard to lab personnel and the environment

Examples of BL-1 Agents: E. coli JM109, DH5a Saccharomyces cerevisiae BIOSAFETY LEVEL 1 (BL-1)

Standard Work Practices Use mechanical pipetting devices Wash hands frequently Minimize splashes and aerosols Decontaminate work surfaces daily Handle wastes properly Maintain insect and rodent control program BIOSAFETY LEVEL 1 (BL-1)

Personal Protective Equipment (PPE) Lab coat or apron Safety glasses or goggles Gloves as needed BIOSAFETY LEVEL 2 (BL-2) Use BL-2 practices when working with: Agents of moderate potential hazard to personnel and the environment

Examples of BL-2 agents: Human blood or body fluids E. coli 0157:H7 Clostridium botulinum Retroviral vectors Human cells in cell culture BIOSAFETY LEVEL 2 (BL-2)

Requirements Adequate illumination Eyewash facility Negative air pressure Autoclave available Biological safety cabinet (BSC) Lab must be separated from public areas BIOSAFETY LEVEL 2 (BL-2) Standard Work Practices Use mechanical pipetting devices Wash hands frequently Minimize splashes and aerosols Decontaminate work surfaces daily Handle wastes properly Maintain insect and rodent control program BIOSAFETY LEVEL 2 (BL-2) Special Practices Place used slides and coverslips in sharps containers, never in any other receptacle. Sharps containers are: • Red in color • Marked with the biohazard symbol • Puncture resistant • Leak proof BIOSAFETY LEVEL 2 (BL-2) Personal Protective Equipment (PPE) • Lab coat or apron • Safety glasses or goggles • Gloves • Biosafety cabinet CASE STUDY CASE STUDY

Opinion in favor of Dr Harran: 1. Why should he be responsible? Most professors don’t provide their graduate students much safety advice anyway. 2. Sangji should have worn a lab coat without the instruction of her advisor, it’s one of the basics of Organic chemistry and she is a graduate student not an undergraduate. 3. Sangji should have read the MSDS for tert butyl lithium, which states that she should not use a plastic syringeand should use a separate glass syringe for each transfer. CASE STUDY

Opinion against Dr Harran: 1. Sangji did not have enough experience to perform the experiment on her own. 2. Dr Harran’s lab did not conform to safety codes. 3. Dr Harran should have advised Sangji to read the material safety data sheets for the chemicals she was using. LESSONS LEARNED

Importance of safety audits and inspections by specialized personnel.

Changing the culture, i.e. The system should help enforce safety by empowering students rather than dictating it from on high. BIOSAFETY CABINETS (BSC)

Biosafety cabinets (BSC) control airborne contaminants during work with infectious material through the use of laminar airflow and high efficiency particulate air (HEPA) filtration. BIOSAFETY CABINETS (BSC)

It is designed to protect personnel and the environment from infectious materials inside the cabinet and to protect the material inside the cabinet from contamination from the lab environment. BIOSAFETY CABINETS (BSC)

Provide product, personal and environmental protection. Various classes of BSCs are available • Amount of air exhausted • Amount of employee protection BIOSAFETY CABINET (CLASS IIB2) http://oregonstate.edu/ehs/LVSG-BSC- • type BIOSAFETY CABINETS (BSCS) Sterile air is exhausted from BSCs through a HEPA filter. • HEPA filter can trap particles to 0.3 microns • Chemical vapors and fumes pass through HEPA filters BSC LOCATION IN THE LABORATORY Since the air curtain created at the front of the cabinet can be easily disrupted, a BSC should be located away from air supply registers, entrances, high traffic areas, and laboratory equipment, e.g. centrifuges, that create turbulence. SAFE AND EFFECTIVE USE OF THE BSC Before beginning work: a.Monitor alarms, pressure gauges, or flow indicators for any changes. b.Turn the cabinet on and let it run for 3-5 minutes. c.Wipe work surface with an appropriate disinfectant, e.g. 70% ethanol. d.Plan your work and place everything needed for the procedure inside the BSC. e.Wipe items with disinfectant before placing in BSC. SAFE AND EFFECTIVE USE OF THE BSC Avoid airflow disruption that could affect the level of protection provided by the BSC: a. Keep the BSC free of clutter, e.g. extra equipment and supplies b. Don’t place objects over the front air intake grille. c. Don’t block the rear air intake grille. d. Limit traffic in the area when the BSC is in use e. Make sure lab door is closed, and avoid opening and closing door if located near the BSC. f. Move arms slowly when removing or introducing items. g. Keep all materials at least 4 inches inside the sash. h. Don’t operate a Bunsen burner in the cabinet. SAFE AND EFFECTIVE USE OF THE BSC

While working: a.Work as far to the back of the BSC workspace as possible. b.Segregate contaminated and clean items. Work from “clean to dirty.” c.Clean up all spills in the cabinet immediately. Allow cabinet to run for 3-5 minutes before resuming work. SAFE AND EFFECTIVE USE OF THE BSC

After completing work: a. Wipe down all items with an appropriate disinfectant before removing. b. Remove all materials and wipe all interior surfaces with an appropriate disinfectant, e.g. 70% ethanol. c. Periodically decontaminate under work grilles. BIOSAFETY CABINETS (BSCS) Helpful Hints . Enter straight into the cabinet, avoid sweeping motions . Don’t place materials on the grill . Keep discard pan or bag inside the cabinet CASE STUDY

Lessons learned: 1. Never add any water to an unmarked container. 2. Label everything with the common chemical name. 3. Always attach your waste label to the waste bottle before you add the waste and fill it out completely and accurately. PROTECTIVE PERSONAL EQUIPMENT (PPE) CONTAMINATION ROUTES

•Ocular invasion

•Inhalation

•Ingestion

•Skin penetration EQUIPMENT PERSONAL PROTECTIVE Head Protection Hearing Protective Clothing Eye, Face,Hand, Foot Respiratory Protection The Strategyof Last Resort

•37 The Occupational Safety and Health Administration (OSHA) requires that personal protective equipment (PPE) be selected, applies to work done with animals too Eye/face/mucous membrane protection When working with potentially infectious materials outside of a biological safety cabinet mucous membranes must be protected by wearing a surgical-type with safety glasses Foot/skin protection Open toed shoes, sandals and other open footwear is prohibited. Shorts are forbidden in laboratories using potentially infectious materials. Protective lab coats are to be worn while in the laboratory. This protective clothing is removed and left in the laboratory before leaving for non-laboratory areas (e.g., cafeterias, library and administrative offices.) PERSONAL PROTECTIVE EQUIPMENT Hand protection GLOVES ARE NEVER A SUBSTITUTE FOR THOROUGH HAND WASHING WHEN ENTERING THE LAB, AFTER REMOVING GLOVES, AND BEFORE LEAVING THE LAB. WASH YOUR HANDS FOR A MINIMUM OF 30-60 SECONDS WITH COPIOUS AMOUNTS OF LIQUID SOAP AND WATER.

OTHER REMINDERS: - Decontaminate work surfaces daily and after a spill involving any viable material. - Properly autoclave your waste. Do not allow waste to accumulate. PERSONAL PROTECTIVE EQUIPMENT BSL 1 and 2 . laboratory coat . gloves . goggles or face shield . masks . hearing protection GLOVE SELECTION CHEMICAL COMPATIBILITY http://www.ehs.okstate.edu/hazmat/gloves.htm www.northsafety.com http://www.showabestglove.com/site/chemrest/ www.ansell-edmont.com/download/Ansell_7thEditionChemicalResistanceGuide.pdf 41

•Laboratory Safety Orientation, 2009 • QUESTION If you wear contact lenses in the laboratory: A. take them out before starting the lab. B. Keep them on but wear protective goggles. C. advise your science instructor that you wear contact lenses. D. keep the information to yourself. QUESTION Personal eyeglasses provide as much protection as A. a face shield. B. safety glasses. C. splashproof chemical goggles. D. none of the above. WASTE DISPOSAL BIOLOGICAL WASTE • Liquid Waste • Sharps • Solid biohazardous waste LIQUID WASTE

• Collect in containers with lids • Autoclave or treat with 10% bleach to sterilize • Dispose of down the drain (as long as no chemical or radioactive contaminants are present) LIQUID WASTE • Procedure: • Collect BL1 and BL2 liquid waste in autoclavable, leak proof containers that are never more than¾full. • Place containers in an autoclavable tray in the autoclave. LOOSEN each container top and place indicator tape on each top. • Adequate cycle time varies depending on load, type of autoclave, and secondary containment. Based on spore testing, determine the • Appropriate cycle time to sterilize liquid waste for your autoclave. Typical cycle times for sterilizing liquid waste range from 45 to 90 minutes at 250°F. • Autoclave temperature should be 250°F (121°C) and autoclave pressure should be 15 psi. • Pour sterilized liquid waste down the sink and flush the drain with water. SHARPS Sharps (needles, syringes, scalpel blades, slides, blood vials, pasteur pipettes) •Collect in approved sharps container •Autoclave to sterilize •Dispose of in medical waste boxes SOLID BIOHAZARDOUS WASTE • Collect BL1 and BL2 waste in a red bag- lined Stericycle biowaste box or reusable bin for pick-up and off-site treatment. The inner red bag must be tied closed. • If using cardboard boxes, close the box with tape and write your building and room number on the side of the box. If liquid is placed in the Stericycle boxes or reusable bins, then place enough absorbent material in the box to absorb the liquid to prevent leaking. • Biowaste boxes or reusable bins are picked-up for off-site Electro Thermal Deactivation (ETD) SAFE WORK PRACTICES SAFE WORK PRACTICES

•WHAT SHOULD IDEALLY BE DONE

•WHAT WE CAN ACTUALLY DO WORK PRACTICES

Education • Hazard recognition and control methods. Training • Proper techniques; emergency response

Supervision- good safety performance BIOLOGICAL SAFETY PROGRAM

The biological safety program of the institution aims at: •Promoting a safe and environmentally-friendly research environment. •Developing institutional and lab specific policies on the safe handling of biohazardous materials, biosafety levels, and personal protective equipment requirements. •Providing training, risk assessment consultations, and laboratory safety audits. •Assisting scientists in the adherence to all federal, state, and local requirements for biological research. BIOLOGICAL SAFETY PROGRAM

PRIORITIES ARE: •Preventing laboratory acquired infections (LAIs) •Facilitating research compliance •Protecting the environment •Raising awareness on the importance of laboratory safety. GUIDING PRINCIPLES 1. Management Commitment and Involvement of Faculty, Staff, and Students 2. Management Responsibility for Safety and the Environment 3. Establishing Clear Roles and Responsibilities 4. Identification of Safety and Environmental Standards and Requirements 5. Obtaining Authorization Prior to Conducting an Activity ADMINISTRATIVE RESPONSIBILITIES Hazard Identification Medical • • in notifications,etc. Signs - Surveillance andimmunization.

•56 MEDICAL SURVEILLANCE MEDICAL SURVEILLANCE

A medical surveillance program is provided through University Health Services (UHS) for personnel who are occupationally at risk of exposure to blood borne pathogens (BBP). UHS provides medical treatment and post- exposure evaluation after an exposure to infectious agents or rDNA for students and employees working in BL1 or BL2 laboratories. e.g. free hepatitis B vaccine offer and post- exposure evaluation with follow up. MEDICAL SURVEILLANCE

A medical surveillance program is also available for personnel that have direct contact with research animals The program includes disease-specific occupational health advice, Animal Biosafety Level 2 training, vaccination offers, pre-screening questionnaires, and post-exposure medical treatment. MEDICAL SURVEILLANCE In addition to being offered recommended vaccinations, BL3 laboratory personnel may have baseline serum samples collected, as appropriate, and tests for agents handled in the lab, e.g. TB skin test.

•NOT APPLICABLE IN OUR INSTITUTION SAFETY AUDITS9 INSPECTIONS

Labs inspected for compliance with • OSHA Lab Standard • Chemical Hygiene Plan • Hazardous, Medical and Radioactive Waste • Labeling, Storage & Segregation • Radioactive isotope use • Biohazardous materials use • Electrical Code SAFETY AUDITS9 INSPECTIONS The Harvad Biosafety office must: Conduct lab inspections Develop emergency and reporting procedures Investigate lab accidents Report incidents and violations of the guidelines. Provide biosafety training. LABORATORY BIOSAFETY AUDITS SUGGESTED IN OUR INSTITUTION

 Assessing appropriate personal protective equipment  Proper use of laboratory equipment (biological safety cabinets, centrifuge, etc)  Proper labeling of equipment for the storing/manipulation of biological materials  Proper procedures for decontamination of biological waste (solid, liquid, sharps)  Presence of life safety devices (eyewash, safety shower, antimicrobial soap, etc) AUDIT PROCESS

Contact department and/or lab representative • Schedule audits for that dept/lab Occupants present • Yes – can ask more – takes longer – better result • No – limited to physical conditions only Preparation of reports 5 Steps of Integrated Safety and Environment Management System (ISEM) (CORE FUNCTIONS) 1. Define scope of activity 2. Identify & analyze hazards 3. Develop & implement controls 4. Perform activity within controls 5. Provide feedback & make improvements QUESTION Your laboratory specializes in virology. There is an epidemic of H5N1 in your country and the Ministry of Health names your laboratory as the reference laboratory for processing all samples. What are the elements to consider ensuring biosafety in your laboratory? EMERGENCY PREPAREDNESS EMERGENCY PREPAREDNESS

•Individual preparedness •Common hazards •Emergency procedures •Emergency communications •Emergency resources EMERGENCY PREPAREDNESS

The primary areas of focus include: •Inclement weather •Natural disasters •Chemical or biohazardous disasters •Fires and floods •Suspicious behavior/activity WHAT TO DO IN THE EVENT OF AN INJURY OR POTENTIAL EXPOSURE: HELPFUL RESOURCES

The Biosafety in Microbiological and Biomedical Laboratories (BMBL) is a CDC/NIH publication that became the cornerstone of biosafety practices and policies in the United States The NIH Guidelines for Research Involving Recombinant or Synthetic DNA The OSHA Bloodborne Pathogen Standard WHO Biological Safety Manual