Definition: Microbes are defined as microscopic organisms agents including bacteria, archaea, protists, fungi including molds, parasites, viruses and viroids. Microbes can be pathogenic (disease causing).
ALL projects involving research with microbes must be pre-approved by the Los Angeles County Science Review Committee (SRC) before experimentation is begun.
Prohibited Research
- Studies involving pathogenic or potentially pathogenic agents are prohibited from being conducted in a home environment. In other words, microbes may not be grown or cultured at home. These types of studies can only be conducted under the direct supervision of an experienced biomedical scientist or designated adult supervisor in an institutional laboratory, including a school, if facilities are adequate and appropriate.
Though cultures may not be grown at home, students MAY collect specimens at home. - Laboratory studies utilizing MRSA (Methicillin-resistant Staphylococcus aureus) and VRE (Vancomycin-Resistant Enterococcus) are prohibited. Microbes KNOWN to be pathogenic are not to be cultured. Pure cultures of non-pathogenic microorganisms should be used in experiments.
- Studies intended to produce microbes with multiple antibiotic resistance are prohibited.
- Microbial studies must be conducted in a properly equipped school or institutional laboratory under qualified adult supervision. No experimentation using existing antibiotic-resistant microorganisms may be conducted unless strains are auxotrophic as well. Auxotrophic microbes cannot survive unless they’re grown on a special medium which contains essential amino acids that they are incapable of producing. They die anywhere outside the Petri dish. Lab Kits for AP Science courses from district-approved Scientific Supply Houses only provide bacterial strains that are auxotrophic.
- Students may NOT be directly involved in the obtaining of microbes (exception: microbe collection in the environment using sterile swabs and appropriate collection techniques and supervision.
Safety Precautions
These safety precautions are intended for experimental activities involving any microbes. Even nonpathogenic microbes may cause disease if they enter the body accidentally. Autoclave or disinfect all waste material; disinfect work areas with 10% bleach, use gloves and goggles.
- Research involving pathogenic or potentially pathogenic agents shall be conducted following standard microbiological practices as defined in Biosafety in Microbiological and Biomedical Laboratories (BMBL) published by CDC-NIH.
- All projects must conform to the CA Education Code Title 2, Division 2, Part 28, Chapter 4, Article 5, 51540.
Regulations and Supervision
The student and Designated Adult Supervisor may consult with the Biomedical Scientist (if required) for detailed guidance in the techniques to be used by the student under the direct continuous supervision of the Designated Adult Supervisor. The Biomedical Scientist or Designated Adult Supervisor must be in the same locality as the student for the length of the experimental work.
The following are examples of precautions that must be taken to prevent injury to persons or the environment. No list could possibly foresee all possible hazards, so teachers, parents and students must carefully plan and follow safe procedures specific to each study. The methods and materials section of the project description must contain explicit and detailed statements as to how and where experiments will be conducted.
- All cultures in Petri dishes must be bound together with transparent tape, immediately after exposure/inoculation. Any Petri dish that contains fungus/mold should be taped shut all the way around the edges. Examine through lids only.
- Inoculating loops must be used with care. Wire loops used for transferring bacteria cultures should be flamed until the entire wire is red hot before and after each transfer is made. Petri dishes that are inoculated with materials containing unknown microorganisms (i.e the material might not be a pure non-pathogenic culture) must not contain blood agar or Brain Heart Infusion (BHI) Broth but rather nutrient or trypticase soy agar.
- These safety precautions are intended for experimental activities involving any bacteria or fungi. Even nonpathogenic microbes may cause disease if they enter the body accidentally. Autoclave or disinfect all waste material; disinfect work areas with 10% bleach, use gloves and goggles.
Glass Petri dishes: to sterilize plates before cleaning or disposal, follow these steps:
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- Autoclave the unopened plates in the usual manner. Usually, steaming at a pressure of 15 pounds per square inch for 15 to 20 minutes kills most microbes. However, to sterilize soil samples or large volumes of culture, continue with the procedure described below.
- Wait one day for any resistant spores to leave the resting stage and begin to grow, sterilize a second time.
- Wait one more day, sterilize a third time — discard sterilized cultures in the regular trash.Disposable plastic petri dishes: place unopened, sealed dishes in Biohazard disposal bags (included in Science Supply kits for E.coli and molds) and use District pick-up of bags as hazardous waste. Calls to nearby universities and hospitals can also yield a place to dispose of microbial waste.All cultured materials must be autoclaved at the end of experimentation according to the recommended procedures in the Science Safety Handbook for CA Public Schools (2014 edition). Exception: Cultures of pure, non-pathogenic bacteria grown in plastic Petri dishes (usually obtained through Science Supply Company kits) can be covered with a 10% bleach solution and allowed to soak for at least 1 to 2 hours. Discard in the regular trash.Sterilizing plates of pure, non-pathogenic bacterial cultures: the materials can be covered with a 10% bleach solution and allowed to soak for at least 1 to 2 hours. Discard sterilized cultures in the regular trash.
Operation of Pressure Cooker (instead of an autoclave) for Sterilization:
- Before using a pressure cooker, the teacher should be familiar with directions for its operation.
- The safety valve should be examined to make sure it is in working order.
- The gauge pressure should be kept at or below a maximum of 20 pounds per square inch.
- The pressure should be returned to zero before the cover can be safely removed.
- The test stopcock should be opened before the clamp can be safely released.
- An eye-protective device should be used when working with a pressure cooker.
4. Experimentation with molds or other fungi must take place in a fume hood or open-air area (to prevent contamination of living areas with fungal spores or exposure to allergens). If anyone in the area has a damaged immune system or any allergies, experiments with molds/fungi must be conducted in a laboratory. Containers must be sealed airtight at all times during observations and disposed of as possible pathogens.
Waste Disposal
Experimentation must include proper disposal methods for the chemicals/materials used in an experiment. The CA Science Safety Handbook describes safe waste disposal for substances used in the lab.
The Flinn Scientific Catalog provides good information for the proper disposal of chemicals. If applicable, the student researcher must incorporate in the research plan disposal procedure required by federal and state guidelines.
Below are the resources needed for Research involving Microbes:
- Microbes Online Pre-Approval template (This is a fillable PDF. Please download this document to your device and enter your responses in the given boxes. You do not need to complete the form in one sitting. Choose Save to save your progress. Once you complete the form, get it approved by your adult supervisor and site coordinator BEFORE you go online to complete the online pre-approval form. You can then copy and paste your responses in the appropriate sections of the online form.)
In addition, Senior Division students need to complete the ISEF Human Informed Consent form and bring them to the fair.