Biosafety Levels in the Laboratory

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Biological research or production labs may work with potentially harmful biological agents which can cause disease in people. These can include pathogens, such as viruses, bacteria, fungi or parasites, as well as some toxic substances. Labs designed to work with hazardous biological agents must have containment features and safe work practices in place to protect laboratory personnel who work with these agents, and to prevent these agents from escaping the lab and harming the community or the environment. These precautions and practices are organized into biosafety levels based on the risks associated with the biological agents being used and the work being done in the lab.

In the United States, biosafety levels are defined by the Centers for Disease Control (CDC) and the National Institutes of Health (NIH). They outline four biosafety levels, or BSLs, that are ranked from 1 to 4 based on the level of risk. BSL-1 precautions are used for the least hazardous biological agents and practices, while BSL-4 precautions are for the most hazardous. BSLs are chosen based on considerations such as what the biological agent is, how much of it is being used, how severe infection could be, how easily the agent could be transmitted, the availability of prevention or treatment for exposure to the agent, the level of exposure risk created by laboratory processes, and worker training and skill levels.

Biosafety Level-1

Biosafety level one is the lowest level of precautions. BSL-1 practices are used for work with agents that pose a minimal risk to workers or the environment and do not typically cause disease in healthy adults. Common examples of agents used in BSL-1 laboratory environments are non-pathogenic strains of E. coli and Bacillus subtillis. While these organisms do present some small amount of risk, standard microbiological practices are usually enough to protect people from them.

Standard microbiological practices include:

  • Hand washing
  • Standard personal protective equipment (gloves, lab coat or gown, and eye protection)
  • Biohazard signs
  • Controlled access to the facility when infectious agents are present
  • Shielding from splashes or aerosols
  • Decontaminating surfaces and equipment at least daily
  • Immediate cleanup and decontamination of spills
  • Safe handling of sharps
  • Prohibiting mouth pipetting
  • Prohibiting food, drink and smoking materials in the lab
  • Decontamination of infectious materials prior to disposal

BSL-1 facilities do not need special containment equipment. Work typically takes place on open bench tops, and the facility does not need to be isolated from surrounding facilities. Because these labs are relatively safe and easy to maintain, they can be used as teaching spaces for workers and students with low levels of training, such as high school biology classes.

Biosafety Level-2

Biosafety level two is for moderate biological hazards. BSL-2 practices can be used for work involving agents that are associated with human diseases (pathogenic or infectious organisms) that pose a moderate hazard to personnel and the environment, such as HIV and the bacteria that cause staph infections. Work that involves human blood or cell lines is also considered a minimum of BSL-2. In addition to standard microbiological practices, BSL-2 laboratories must follow these additional practices:

  • Lab personnel are trained to handle pathogenic agents and are supervised by scientists with advanced training
  • Biosafety cabinets or other physical containment for all procedures that can generate infectious aerosols or splashes
  • Autoclave or other method of decontamination for proper disposal
  • Eyewash station
  • Lockable doors and more controlled facility access
  • Extra care is taken to control routes of exposure, including advanced techniques for handling contaminated sharps
  • Immunizations are provided to lab personnel when appropriate
  • Additional PPE, such as face shields, may be necessary
  • A lab-specific biosafety manual that outlines the necessary controls and practices for the work performed in that lab

BSL-2 facilities will need moderate containment methods such as Class II biosafety cabinets. All surfaces in the lab must be easy to clean and decontaminate effectively. Carpets and rugs are not allowed, and windows must be fitted with screens.

Biosafety Level-3

Biosafety level three is for serious biological hazards. BSL-3 practices are appropriate for work involving agents that can cause serious or potentially fatal disease through inhalation. Examples of agents commonly used in BSL-3 work include yellow fever virus, SARS coronavirus, and tuberculosis bacteria. BSL-3 work is often strictly controlled by government agencies, and labs may need to be registered.

BSL-3 labs must follow all of the same practices as BSL-1 and BSL-2 labs. In addition, BSL-3 labs must incorporate stricter measures including:

  • Baseline medical testing and ongoing medical surveillance for all workers with potential exposure to infectious agents
  • Full body PPE such as wraparound gowns, scrub suits or coveralls
  • Respiratory protection may be required
  • All work with infectious agents must be performed in an appropriate BSC or other physical containment device
  • Access is restricted and controlled at all times
  • More stringent control of contaminated waste, equipment and lab clothing
  • A set of two separate, self-closing doors, separated from general building corridors

BSL-3 facilities must have more advanced containment methods, including specialized ventilation that directs air from clean areas towards areas where infectious agents are present, and does not allow air to recirculate unless it runs through a HEPA filter first. Windows must be sealed so that airborne particles cannot escape.

Biosafety Level-4

Biosafety level four is the highest level of precautions. BSL-4 practices are used for work with agents that are very easily transmitted and cause serious or fatal diseases for which there are no vaccines or treatments, such as the Ebola virus and the virus that causes smallpox. These facilities are rare and are highly regulated.

In addition to the precautions used in BSL-1, 2 and 3 facilities, BSL-4 labs must use practices such as:

  • A complete clothing change before entering the lab, and a decontamination shower before exiting
  • Decontamination of all materials before leaving the facility
  • Strictly-controlled access and records of all persons entering and exiting the facility
  • An airlock entrance
  • Perform all work in a class III biosafety cabinet, or a combination of a class I or II biosafety cabinet and a positive-pressure full-body suit with an air-supplied respirator

BSL-4 work must typically take place in a dedicated building or a completely isolated area of the facility with dedicated air intake and exhaust, and dedicated vacuum lines and decontamination systems. Air exhaust and used water must pass through HEPA filtration before leaving the facility.

More information about biosafety levels and containment practices can be found in the CDC publication, Biosafety in Microbiological and Biomedical Laboratories, 5th Edition.

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