Concepts of Biocontainment & Biosafety Training UNIVERSITY OF NORTH CAROLINA - CHARLOTTE/ APRIL 30, 2014 CLIENT NAME / DATE Regulatory Guidance . OSHA Bloodborne Pathogen Standard . NIH Recombinant Guidelines – Apply to all institutions receiving NIH funding – Covers rDNA work, but also includes risk group listing . Biosafety in Microbiological and Biomedical Laboratories (BMBL) . Select Agent and Toxin regulations . Dangerous goods shipping regulations . North Carolina Dept. of Environment and Natural Resources CLIENT NAME / DATE US Guidance for Laboratories . BMBL Outlines: – Standard Microbiological Practices – Special Microbiological Practices – Safety Equipment – Facilities . Includes – Agent summary statements – BSC design and use – Toxin handling http://www.cdc.gov/biosafety/publications/bmbl5/index.htm CLIENT NAME / DATE Layers of Protection CLIENT NAME / DATE Definitions . Biohazard – A biological agent capable of self- replication that can cause disease in humans, animals, or plants – Generally, a microorganism, or toxins and allergens derived from those organisms . Biocontainment Class III cabinets at the U.S. Biological Warfare Laboratories, Camp Detrick, Maryland (Photo, 1940s) – the physical containment of pathogenic organisms or agents to prevent accidental infection of workers or release into the surrounding community . CLIENT NAME / DATE Biosafety . Fundamental objective: – Containment of potentially harmful biological agents . Risk assessment: – Helps to assign the biosafety level that reduces to an absolute minimum the worker’s exposure to agents, their risk of an LAI (lab associated infection), and potential impact on the community and the environment CLIENT NAME / DATE Biosafety Levels (BSLs) . Combinations of lab practices and techniques, safety equipment, and laboratory facilities . Appropriate level determined by risk assessment . Specifically appropriate for: – Organism in use – Operations performed in the laboratory – Known or suspected routes of infection CLIENT NAME / DATE Determining BSLs . Conditions under which the agent ordinarily can be handled in the laboratory . Final determination by responsible scientists on the Institutional Biosafety Committee (IBC) . Based on a risk assessment – Resources include: . Published data, including animal data . Epidemiological data . Guidelines and regulations . Biological safety officer (BSO) CLIENT NAME / DATE RISK ASSESSMENT PROCESS CLIENT NAME / DATE Basic Sources of Information . The NIH Guidelines for Research Involving Recombinant or Synthetic Nucleic Acid Molecules . Risk Group listing . Public Health Agency of Canada . Pathogen Safety Data Sheets and Risk Assessment . CDC Agent Summary Statements – In BMBL . CDC website . CDC/MMWR recommendations . Others CLIENT NAME / DATE Risk Assessment . Consideration of: – Manipulations to be done – Volumes handled – Virulence – Pathogenicity – Antibiotic resistance patterns – Vaccine and treatment availability – Other factors CLIENT NAME / DATE Risk Management— Remember the “Chain of Infection” Presence of a pathogen Susceptible Pathogen is host virulent Sufficient Portal of numbers of entry organisms CLIENT NAME / DATE Infectious Dose . Agent . Dose • Ebola virus • 1 Salmonella, NIH • TB • 1 - 10 • Tularemia • 10 • Anthrax (inhalational) • 8,000-50,000 est. • Cholera • 108 • Salmonella typhi • 105 • E. coli (enteropathogenic) • 108 – 1010 • Shigella • 10 – 200 • Norovirus • 18 CLIENT NAME / DATE Routes of Exposure . Parenteral (needlestick, scratch) . Exposure to non-intact skin . Inhalation . Droplet . Ingestion . Mucous membranes . Absorption (e.g., toxins) . Animal bites and scratches CLIENT NAME / DATE Laboratory Associated Infections (LAIs) . Where no known exposure event – usually attributed to aerosols . Aerosols can be produced by equipment: – Centrifuges – Blenders – Sonicators – Vortex mixer – Homogenizers, etc. Aerosols also produced by: – Any liquid manipulation – Pipetting practices – Opening lyophilized cultures – Flaming loops and needles – Changing bedding of infected animals CLIENT NAME / DATE Aerosols . Lab equipment – Produces aerosols more efficiently than natural methods (coughing, sneezing, etc.) . Materials encountered – Higher titered than in natural setting . Modest aerosol-producing activity could be infectious – Larger quantities being manipulated – Agent that is not normally aerosol-transmissible may be transmitted under these circumstances . Use the BSC to contain aerosols – Class I or II BSC for containment of small equipment (blenders, homogenizers, etc.) Banthrax Versa Dome CLIENT NAME / DATE Particle Size and Routes of Transmission . Large droplets (>50-100 µm in diameter) – Subject to gravity – Travel only a few feet – Do not remain suspended for very long . Intermediate droplets (10-50 µm) – Affected by temp, humidity, air velocity and air currents – With water loss, may become “droplet nuclei” quickly . Small particles (<10 µm) – Remain airborne for extended periods of time – Dispersion affected by air currents – Infective if organism can survive desiccation – Can reach the pulmonary region of the lung with variable efficiency CLIENT NAME / DATE Particle Size and Routes of Transmission CLIENT NAME / DATE Susceptibility to Infection . Genetics . Inherited immune problems . “Healthy Adult” . Factors affecting immune status: . Chemotherapy, smoking, immune suppressive drugs, . Lupus, HIV, etc. CDC/James Gathany . Immune function usually decreases with age . Vaccination Status . Reduces chance of infection . Reduces severity of infection (chickenpox) . Never reduce risk 100% . Reproductive Risks . Risks to fetus: Toxoplasma, Listeria, CMV, Rubella, HIV . Mother may be asymptomatic, but fetal effects are severe CLIENT NAME / DATE HIERARCHY OF CONTROLS CLIENT NAME / DATE Hierarchy of Controls CLIENT NAME / DATE CONCEPTS OF BIOCONTAINMENT LABS (ENGINEERING CONTROLS) CLIENT NAME / DATE Biocontainment Concepts . Containment Barriers – Primary - BSC’s, personnel protective gear, containment equipment – Secondary - Room, systems – Tertiary - Containment around systems . Access Control and Separation . Redundancy and Reliability . Decontamination CLIENT NAME / DATE Biosafety Levels 1 & 2 BSL-1 BSL-2 CLIENT NAME / DATE Directional Airflow Provides “Zone Control” of hazards and odors Air flow from lowest to highest hazard CLIENT NAME / DATE BSL-3 CLIENT NAME / DATE BSL-3 facility design . Isolated from other lab areas . Access through two self-closing doors . Single pass, unidirectional air in-flow . HEPA filters not required on exhaust air . Interior surfaces sealed to allow decon . Air space around penetrations sealed . Autoclave available in facility CLIENT NAME / DATE BSL-4 Labs . Suitable for work with dangerous and exotic agents that pose a high individual risk of aerosol-transmitted laboratory infections and life-threatening disease. .Exposure potential to pathogens – That may be spread by aerosol or – With unknown risk of transmission . Infection possibly lethal . Examples: . Ebola Zaire . Sin Nombre virus . Rift Valley Fever CLIENT NAME / DATE Biosafety Level 1, 2, 3 . Provide access to autoclaves to treat potentially contaminated waste materials (e.g., gloves, lab coats, etc.) from laboratories before disposal or reuse CLIENT NAME / DATE All Biosafety Levels . Sinks are required to wash hands after: – Handling infectious materials – Removing gloves before leaving the lab – Spills and/or exposures . Ideally, the hand washing sink is located at the exit to each laboratory . Non-hand operated is recommended CLIENT NAME / DATE Biological Safety Cabinets (BSCs) . Class 2, Type A cabinets . 2 HEPA (high efficiency particulate air) filters . Filtered air discharged to room (no volatile toxic compounds) . Use to contain aerosols and maintain sterility of specimens . Protect you, your work and the environment . Dependent on scrupulous work practices . Read BSC Operating Procedure CLIENT NAME / DATE Vacuum Set-Up CDC drawing Flasks should be plastic, plastic-coated glass or covered with netting A – disinfectant containing flask B – overflow flask C – in-line HEPA filter D – vacuum system CLIENT NAME / DATE ADMINISTRATIVE AND WORK PRACTICE CONTROLS CLIENT NAME / DATE Purpose of Administrative and Work Practice Controls Protection of: . yourself . ”work products" . co-workers . lab support personnel . community . environment CLIENT NAME / DATE Safe Work Practices . Wash hands frequently Fill line . Do not bend or recap needles . Place sharps in the appropriate container . Do not eat, drink, smoke, apply cosmetics or handle contact lenses in the work areas CLIENT NAME / DATE Safe Work Practices . Minimize splashing, spraying, generation of droplets . Do not reach into trash/sharps containers . Minimize glassware/sharps hazards . Never pipette by mouth – use pipet aids . Wipe down work area with a disinfectant CLIENT NAME / DATE Proper Use of the BSC • Work 4” back from the • Load only those materials front grille needed inside cabinet – Avoid withdrawing hands for • Do not place items on the discard or supplies front grille or block rear • Wait 5 minutes after loading vent BSC before working • Wipe all materials going • Avoid rapid sweeping arm in and out of BSC with movements disinfectant (depending) • Clean trough under work surface regularly Place discard containers • • No open flames inside inside BSC cabinet CLIENT NAME / DATE Minimize use of sharps– use alternatives . Hypodermic needles . Use safety needles / syringes . Pasteur pipet . Use plastic transfer pipet . Scissors . Use