Under what conditions can I generate bacterial isolates from environmental samples in a BSL-1 environment?
I have heard of many academic labs (or even undergraduate classes) generating bacterial isolates from environmental samples in a BSL-1 environment. Including SynBERC/JBEI’s iCLEM program, where high schoolers isolate novel cellulose degrading strains from compost samples, and one professor involved who was having his students isolate high-priority unsequenced gut microbiome strains.
My impression was that strictly speaking, this kind of work is not allowed in a BSL-1 environment, and so far we have not been allowed to do anything like this at BioCurious either. Is this correct? Apparently, many people in academia feel otherwise. Under what conditions should we be able to do this safely in a BSL-1 lab?
Answer from a Biosafety Officer:
May 6, 2013
You are correct. According to best practices in biosafety, use of Biosafety Level 2 (BSL-2) practices and facilities is recommended when working with samples that contain unknown microbes, and therefore unknown hazards (See Ref #1 below, Section II: Biological Risk Assessment; 2: Isolation of Unknown Microbes from the Environment).
However, in practice, biosafety is built on a strong foundation of situation-specific risk assessment. For specific, well-described experiments and/or projects with facility-specific information it is possible to develop procedures that can allow some types of experiments to be performed in what is technically a BSL-1 laboratory using BSL-2 work practices. It is difficult to generalize this advice without specific information, but site-specific risk assessments and work practices could explain how the specific experiments that you mentioned in your questions are being performed safely.
Here is a summary of standard good laboratory practices from the CDC/NIH publication Biosafety in Microbiological and Biomedical Laboratories (BMBL). Standard Good Laboratory Practices for BSL-1 and above is:
- Your work area should be dedicated for work with microbes. Don’t store cultures or plates in your kitchen. Ideally, cultures should not be stored in living areas such as bedrooms.
- Always wash your hands after working with microbes and before leaving your work area.
- Do not eat, drink, apply cosmetics, or store food or toiletries in your work area.
- Don’t mouth pipet.
- Avoid the use of needles, scalpels, and other sharp items with infectious microbes if possible. Make sure that you dispose of these items properly – sharps containers and return programs available at some pharmacies.
- Use careful work practices to avoid splashes and production of aerosols.
- Decontaminate your work area when you are finished working. A simple mixture of 1 part Clorox bleach to 9 parts water is a very effective disinfectant. Make sure that all surfaces that you want to disinfect remain wet with the diluted bleach solution for at least 5-10 Minutes.
- Decontaminate your cultures and plates before disposal. Cultures should be treated with a 1:10 dilution of Clorox bleach (1 part bleach to 9 parts liquid culture e.g. 10 mL bleach in 90 mL liquid culture). The surfaces of petri dishes can be soaked in a 1:10 dilution of Clorox bleach for 2 hours before disposal (2).
- Wear a lab coat to prevent contamination of your clothing.
- Wear gloves to protect yourself from exposure to infectious agents. Change your gloves if they become torn or contaminated. Remove your gloves when you are finished working and wash your hands before leaving your work area. Don’t wash or reuse disposable gloves. (1)
Additional Good Work Practices for BSL-2:
- Any work with infectious agents that may result in the formation of aerosols should be done inside of a biosafety cabinet (BSC) or other containment equipment.
- Lab workers should be offered medical services and immunizations if they are available for the infectious agents that are handled in the laboratory.
- A biosafety manual describing laboratory-specific procedures for safe conduct of experiments, decontamination of cultures, waste handling, spill clean up, decontamination of surfaces, procedures to be followed in case someone is exposed to an infectious agent, etc.
- A supervisor should make sure that laboratory members demonstrate proficiency in standard and special practices before working with infectious agents.
- Durable, leak-proof containers must be available to contain potentially infectious materials while collecting, handling, processing, storing, or transporting in the laboratory.
- All equipment should be decontaminated on a routine basis, as well as after any known spills, splashes, or other contamination.
- Spills of infectious agents must be contained, decontaminated and cleaned up by trained personnel with appropriate equipment.
- All equipment must be decontaminated before repair, maintenance, or removal from the laboratory.
- Potential exposures to infectious agents must be immediately evaluated and treated by medical personnel. Incident response procedures should be described in the laboratory’s biosafety manual. Medical evaluation, initial and follow up testing, and treatment should be provided and records should be maintained.
- There should be no animals or plants in the work area unless they are directly associated with the experiment that is being performed. (1)
The other reference document, Draft Guidelines for Biosafety in Teaching Laboratories, also contains useful information for modification of practices for teaching laboratories (2). For example, autoclaves are the gold standard for decontamination of biological wastes. However, in instances where a laboratory does not have an autoclave, the surfaces of petri dishes can be soaked in a 1:10 dilution of Clorox bleach for 2 hours before disposal (2).
The most important limiting safety feature for most laboratory facilities is whether or not you have access to a biosafety cabinet. When working with environmental, human, or other unknown samples best practice is for any manipulation of bacterial isolates to be performed in a biosafety cabinet.
1. Wilson, D., and L. Chosewood, editors. Biosafety in Microbiological and Biomedical Laboratories (BMBL), 5th edition. Centers for Disease Control and Prevention and National Institutes of Health. 2009. http://www.cdc.gov/biosafety/publications/bmbl5/ Accessed 03/03/2013.
2. American Society for Microbiology. Draft Guidelines for Biosafety in Teaching Laboratories. 2012. http://www.asmcue.org/documents/ASMBiosafetyGuidelines-v2.pdf Accessed 03/03/2013.