Issues relating to culturing of microorganisms
Background
Microorganisms are microscopic organisms, such as bacteria, fungi (moulds and yeast), viruses, microscopic algae and protozoa.
Risk group 1 organisms, as described in AS/NZS 2243.3:2010, are generally regarded as suitable for supervised school experiments, if the laboratory complies with the requirements for Physical Containment Level 1 (PC1). Different school authorities have different views on the culturing of microorganisms and the disposal of biological wastes. Please check with your school authority for details.
Experiments should take into account such factors as skill level and behaviour of students, class size, available facilities and the level of training and experience of staff.
Culturing microorganisms can lead to the growth of dangerous pathogens. Pathogens may enter the human body through skin, eyes, puncture wounds, inhalation, or ingestion. Students who have compromised immune systems should consult the teacher or their doctor before participating in microbiology experiments. Risks are minimised by:
- Careful choice of organisms or sources of organisms from the environment, so as to avoid culturing pathogens. Pure non-pathogenic strains of organisms can be ordered from reputable suppliers. Many school authorities recommend using only microorganisms obtained from known sources.
- Culturing in closed containers, which are taped before examination and remain unopened, so that there is no generation of harmful microbial aerosols.
- Staff being specifically trained in microbiology and aseptic techniques, including the minimisation of the production of microbial aerosols.
- No sub-culturing or transferring of organisms from one medium to another (unless appropriately trained in microbiology), since this might concentrate pathogens.
- Disposing of all waste material appropriately, ensuring that all microorganisms are killed before disposal.
Some possible ways to reduce risks:
- Do not collect microorganisms from human body fluids (including from sneezes or coughs) or skin (other than hands and fingers), or from toilets, since this reduces the likelihood of culturing pathogens.
- Do not culture from animal sources (e.g bird cages, dead animals, faecal material) to reduce the likelihood of culturing pathogens.
- Do not use agar culture media that selects for human pathogens e.g. blood agar, MacConkey's agar, dung or faecal agar.
- Students should expose growth media to the environment for the minimum time. When transferring colonies, the Petri dish cover should be held at 45 degrees to minimise exposure to air.
- Do not incubate plates above 30ºC. Incubation above this temperature may encourage pathogens that grow at human body temperature. Incubators should be disinfected with a chemical disinfectant before and after use.
- Invert agar plates before incubating to avoid condensation dripping on to cultures. Any drips from a partially sealed Petri dish are potential sources of infection.
- Tape the lid and base of each agar plate together with 2-4 short strips of adhesive tape, but not around the full circumference. This reduces the possibility of growing anaerobic organisms that may be pathogenic.
- Ensure the plates are sealed before inspection by students.
- Students should avoid contact with cultures, and introducing objects into their mouths such as eating, licking labels, biting nails or sucking pencils. Contact lenses should not be adjusted or cosmetics applied during the experiment.
- Only essential papers should be kept on the laboratory bench. Personal items, such as mobile phones, calculators, pens, pencils and cosmetics, should not be handled, as this could result in the items being contaminated.
- Do not store food or drinks in a refrigerator where microorganisms are stored.
- All cuts or broken skin of students and staff should be covered by a water resistant dressing and gloves before class. Hands should be washed with soap and water after the gloves are removed. Gloves should be removed using the appropriate techniques so that cross-contamination does not occur.
- Never pipette liquid cultures by mouth.
- All cultures, media, chemicals and disinfectant should be clearly labelled. Hazardous substances should have their hazard information clearly visible.
- Count plates out and in again, if there is any chance of students taking the plates away.
- Students should wash their hands thoroughly with soap and water before the experiment and before leaving the laboratory.
- Ensure that disinfectant solution is available, should there be a spill. All spills should be cleaned up carefully using appropriate techniques. Contaminated objects should be disinfected or disposed of appropriately.
- Disinfect all bench and work areas before and after working with microorganisms. The optimal concentration of ethanol is 70% in water. Ensure that there is no ignition source available if a flammable liquid, such as ethanol or methylated spirits, is used as a disinfectant. Wear safety glasses when using disinfectants, as disinfectants may be dangerous to the eyes. Staff and students should be aware of the location of the eye wash station before beginning the experiment.
- Do not keep the cultures for more than 7 days.
- Sterilise cultures and all waste material by placing them in autoclavable biohazard bags and using an autoclave or pressure cooker (see manufacturer's directions for appropriate time, pressure and temperature). The autoclavable biohazard bag should be sterilised when only half-full, otherwise a spill of contaminated material could occur. Do not place sharps in the autoclavable biohazard bag. Chemical disinfectants may be inactivated by contact with organic material and are not recommended. After sterilization, cultures may be disposed of in the autoclavable biohazard bags in an industrial waste bin.
- Never autoclave any flammable material.