THE UNIVERSITY OF NEWCASTLE- SCHOOL of BIOMEDICAL SCIENCES

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1 Page: 1 of Purpose: 1.1 To list the legal requirements and University Policies pertaining to the use of microorganisms in the Laboratory. 1.2 To outline the guidelines and recommendations for working safely with microorganisms. 2. Equipment: N/A 3. Materials: 3.1 Other General Discipline Procedures as cited. 4. Set Up: N/A 5. Safety Precautions: 5.1 This information has been compiled from the references as listed in section 10. For more information consult these directly. 5.2 Classification of Microorganisms All microorganisms and genetically modified microorganisms are classified into 1 of 4 Risk Groups. The group definitions are as follows: Risk Group 1 (low individual and community risk) - it is unlikely that these microorganisms will cause animal, human or plant disease. Risk Group 2 (moderate individual risk, limited community risk) - these pathogens can cause animal, human and plant disease, but are unlikely to be a serious hazard to laboratory workers, the community, livestock or the environment. If infection does result from laboratory exposure, treatment and preventative measures are readily available and there is a limited risk of the infection spreading. WRITTEN BY CHECKED BY REVIEWED BY AUTHORISED BY NAME (signed) Sarah Cooper Melissa Musicka Lynn Herd Lynn Herd DATE 6th July th July rd February th June 2005 Distributed To: GDP Master file / GDP Lab file

2 Page: 2 of 10 Risk Group 3 (high individual risk, limited community risk) -These pathogens usually cause serious animal or human disease and may present a risk to laboratory workers. They may present a risk if spread in the community, but there are usually effective preventative measures or treatment available. Risk Group 4 (high individual and community risk) - These pathogens usually produce life-threatening animal or human disease. They represent a serious hazard to laboratory workers and are readily transmittable from one individual to another. Effective treatments and preventative measures are not usually available. NB Prior to the commencement of work with any microorganism, determine which Risk Group it belongs to. (See 5.3). 5.3 Classification of Facilities PHYSICAL CONTAINMENT LEVELS (PC) set out certain facility requirements for the use of pathogens. They are numbered from 1 to 4 and correspond to Risk Group numbering. Tables of various pathogens and their Risk Group classifications are listed in Section 3 of the Australian/New Zealand Standard - Safety in laboratories, Part 3: Microbiology (AS/NZS : 2002) This Standard is available from Health and Safety PHYSICAL CONTAINMENT LEVEL 1 (PC1) - No special containment facilities are required for the use of Risk Group 1 microorganisms. They present no risk to healthy adults and workers should be protected from any risk by standard laboratory practice. Work may be carried out on an open bench. (Human blood and body fluids should not be handled in a PC1 facility). General guidelines for microbiological work carried out in a PC1 Laboratory include: (for a complete requirements list, see the Microbiology Standard) A lab coat (preferably a wrap around rear fastening gown) must be worn to protect the front of the body Closed footwear must be worn Protective clothing must be removed before leaving the laboratory. Cover all exposed wounds or cuts (especially on the hands) with waterproof dressing. Safety glasses and face shields must be worn (where appropriate) if there is a risk of splashes or other hazards. (Contact lenses are not a

3 Page: 3 of 10 form of eye protection) Food or drink must not be brought into, consumed or stored in the laboratory. Smoking, shaving and applying cosmetics is prohibited in laboratories. Mouth pipetting is prohibited. Hands and nails must be washed with an antimicrobial soap before leaving the laboratory and moving to areas such as the toilet, tearoom or office. Only wash hands in the designated hand-washing sink. This sink should be kept clean and is not to be used for any other purpose. Be aware that while working in the laboratory, hands, pencils and notebooks may come into contact with contaminated surfaces, aerosols and liquids, and must be kept away from the face. All cultures and samples must be clearly labeled and stored appropriately i.e. refrigerators, incubators. Work carried out on the bench must be done in such a way so that aerosol production is minimized. Long hair must be tied back Significant spills and accidents must be reported. Workbenches must be decontaminated after spills and when work is complete. Use caution when dealing with spore producing fungi. Ensure that petri dishes are sealed with tape and that incubators are allocated for fungi use only. Only use an allocated bench to write up on. This bench should be kept free from contaminated substances and lab work is not to be carried out on this space. Personnel who wish to transport infectious materials between institutions should be aware of regulations that exist in relation to this. See Section 8 of the Australian/New Zealand Standard - Safety in laboratories, Part 3: Microbiology. (AS/NZS : 1995) and GDP 012 Packing Goods For Shipment. All unwanted wastes containing live microorganisms must be sterilised by autoclaving, treated by a chemical disinfectant, or incinerated in an EPA-approved incineration facility; before disposal (before it goes offsite).

4 Page: 4 of PHYSICAL CONTAINMENT LEVEL 2 (PC 2) - is required for any facility in which pathogens from Risk Group 2 are to be used. All facilities and procedures of a PC 1 laboratory apply, plus additional requirements, including (for a full list refer to the Microbiology Standard): Gloves should be worn when handling blood and body fluids or infectious materials. Hands must be thoroughly washed after gloves are removed and discarded with the infectious waste. Gloves must not be reused. Biological safety cabinets must be used where infectious aerosols are likely to be produced. Access to the PC2 laboratory must be restricted only to those who have management authorization. Laboratory doors must be closed when work is in progress. All clinical specimens must be regarded as being potentially infectious. A biological safety cabinet must be used for the sonication, shaking and mixing of Risk Group 2 substances, unless the equipment incorporates an aerosol containment device. At least 5 minutes must elapse before opening any of these devices to allow for the settling of aerosols. Safety glasses and gloves must be worn during the subculture of Risk Group 2 organisms. Vaccinations may also be recommended. Any materials being transported to other areas of the building (including waste) must be enclosed in a second unbreakable container or sealed bag. Laboratory staff must advise any maintenance, service or cleaning personnel of the special microbiological hazards in the laboratory. Cleaners must never be expected to dispose of contaminated waste. Potentially contaminated glassware and instruments must be autoclaved or chemically disinfected prior to washing and re-use. All microbial waste must be incinerated or autoclaved before disposal (before it goes off-site). A pest control system against insects (and other animals) must be instituted. The smelling or sniffing of bacterial cultures for odours is prohibited. Any lab air-conditioning must maintain a negative pressure in the lab by extracting air. Air recirculation is permitted, but not to areas outside the PC2 facility.

5 Page: 5 of PHYSICAL CONTAINMENT LEVEL 3 (PC3) - must be utilized when Risk Group 3 pathogens are used. Work in a PC3 facility includes all the practices and equipment used in PC2 laboratories, plus additional requirements (for a full listing, see the Microbiology Standard); A PC3 laboratory must be physically separated from other areas and not accessible by the general public. A double door system that includes an airlock must be present. Laboratory windows are to be permanently sealed. All equipment for PC3 requirements must be dedicated to that use and area. Sterile liquid effluents must be discharged directly to the sewerage system. Laboratory clothing must be laundered regularly. Clothing used in hazardous areas must be segregated and disinfected or sterilised before cleaning. Personal clothing and effects must be stored in facilities adjacent to the laboratory area and must not be taken into the laboratory. Only staff who have been especially trained and who meet specified requirements are permitted in the laboratory. Where an autoclave is not available within the laboratory, wastes must be bagged and placed in an unbreakable container with a secured lid for transport to the autoclave. The surfaces of the container must be decontaminated with a suitable disinfectant. Wastes must not be stored outside the PC3 laboratory before sterilisation. Lab doors must remain locked whenever the room is not occupied. All procedures involving PC3 infective materials (or when required by the OGTR) must be conducted in a biological safety cabinet, Class I or II. * OGTR is the 'Office of the Gene Technology Regulator'. No one must enter the laboratory for repairs or cleaning until the surfaces have been disinfected and authorisation has been obtained from the laboratory supervisor or safety officer. Protective clothing must not be worn outside the laboratory and must be transported to the decontamination area in sealed bags or boxes that facilitate steam penetration during autoclaving. Personnel should be subjected to an initial medical examination,

6 Page: 6 of 10 (including chest X-ray where appropriate) and periodic examinations. A baseline serum sample should be obtained from staff before commencing work in a PC3 facility. Immunisation should be considered where appropriate. For more information on building specifications and PC4 laboratories, see CCH Laboratory Safety Manual. 5.4 For a list of locations of all University PC2 and PC3 Laboratories see Illustration The University of Newcastle PC2/PC3 Facilities. NB The University does not have any PC4 facilities. 5.5 GENETICALLY MANIPULATED ORGANISMS Anyone planning work with genetically manipulated organisms must consult the current edition of The Office of the Gene Technology Regulator Guidelines. Also see GDP 009 Molecular Biology Safety and the Biological Safety Section of the CCH Laboratory Safety Manual. 5.6 CLINICAL SPECIMENS - refer to GDP 010 Clinical Specimen Safety for specific safety guidelines to be followed when dealing with human specimens/samples. NB Human blood and body fluids must be handled in a PC2 facility or higher. 5.7 WASTE DISPOSAL Waste must be separated at source into contaminated/non contaminated waste, and, if contaminated into reusable equipment and material for disposal. All equipment that has come into contact with material of plant, microbial, animal or human origin is to be handled as if contaminated (i.e. is a potential danger to humans, animals, agriculture or the environment) for the purpose of laboratory manipulation and waste disposal. All contaminated waste materials (including those containing genetically modified organisms) are to be sterilised (preferably by autoclaving) before final disposal (before it goes off-site). Solid contaminated materials are to be placed in autoclavable biohazard waste bags. Contaminated broken glass and sharp objects are to be placed in Sharps Containers for disposal. Culture media or other liquid wastes, which may contain viable organisms, may be discarded into the sewer, but only after sterilisation by autoclave or

7 Page: 7 of 10 chemical means. All sample remains, disposable equipment, animal faeces and bedding, should be regarded as contaminated. Wherever possible, contaminated wastes should be placed in containers which will not entrap air, and be autoclaved before being discarded or incinerated. Aerosol cans and other sealed containers which may explode on autoclaving or incineration should be surface sterilised only before discarding. Animal remains should be bagged, labelled with users name, date, weight and known contamination and stored frozen prior to collection and incineration. The local company, Cleanaway, currently performs this task for the University. (See GDP 015 Animal Handling Procedure for more detailed disposal directions). Reusable glassware etc., contaminated with microorganisms, must be disinfected or autoclaved (preferably both) before cleaning. Containers for contaminated equipment must be readily distinguishable from those used for ordinary dirty glassware etc. Care must be taken to ensure that containers of disinfectants (e.g. for pipettes) are deep enough to permit contact of disinfectant with the whole of the contaminated surfaces. Containers for infected equipment, should, where appropriate, be provided with lids and, if these containers are also used for autoclaving the contents before cleaning, they should be designed to entrap minimal air. Used plastic Petri dishes, culture bottles and tubes for final disposal should be carefully placed in solid-bottomed containers (not wire baskets) or in (double) autocalvable bags. After autoclaving they should be incinerated. Trained personnel must supervise the disposal procedure. Bench areas on which procedures on material of plant, microbial, animal or human origin has taken place should be treated as contaminated and wiped with a 5% solution of sodium hypochlorite at completion of experiments or at the end of each day. 5.8 ACCIDENTS/EMERGENCY PROCEDURES The first concern in an accident should be the care of the person(s) injured or at risk from infection. The First Aid Officer should be called and then medical aid sought in all cases of injury or doubt. Injured persons should not be moved, unless in further danger, e.g. from fire, gases, until expert assurance is obtained that movement will not cause further injury. Where aerosols have been generated, the area should be evacuated, sealed

8 Page: 8 of 10 off and signs put up to prevent people entering the affected area. Due to the recirculating nature of the Air-conditioning system in the LSB, the airconditioning may need to be switched off to prevent aerosols produced by a Microbiological Spill from entering other areas. Maintenance can be contacted to purge or switch off the system. See the 'LSB Safety Notice' attached for details (Attachment 8.2). Allow time for particles to settle before disinfecting all surfaces liable to have been contaminated. Remove protective clothing (if contaminated) and wash hands and arms with antimicrobial soap. Apply clean protective clothing (lab coat, gloves, safety glasses, mask) and proceed with decontamination. Disinfection procedures for spills of infectious material should be rapidly effective and aim at containing the problem in the affected area. Spills, such as knocking over liquid cultures or dropping culture plates, should be treated with a general disinfectant. Pour the solution around the spill, allowing it to mix gradually with the contaminated material. Avoid pouring the disinfectant directly onto the spill, as this can produce more aerosols. Paper towels soaked in disinfectant can be used to cover the area. Allow at least 30 min for it to act. Using the same disinfectant, wipe over surroundings likely to have been contaminated with aerosols. After another 30 min, the spillage and disinfectant should be carefully mopped up and all contaminated objects and liquids transferred to a metal tray for sterilisation in an autoclave. Where a spillage of a specimen from a known or suspected, Hepatitis B, Hepatitis C or HIV patient occurs, a fresh sodium hypochlorite solution (5000ppm) should be used. Waste materials that contain hyperchlorite must not be autoclaved due to the subsequent production of toxic gas. A full report of any accident and measures taken, along with injury report and hazard notification (if applicable) must be made as soon as possible to the Occupational Health and Safety Officer. For more information on Disinfectants and antiseptics see document titled 'The University of Newcastle Occupational Health and Safety Committee - Guidelines for the Safe Use and Disposal of Biological Material and Material of Biological Origin'. Also see GDP 011 Spill Procedure. 6. Maintenance: 6.1 If any part if this GDP changes, attach new information so that they may be

9 Page: 9 of 10 included when an updated issue is produced. 7. Shutdown: 7.1 If you were required to read this GDP, sign the Requested Reading Record Sheet (Illustration 5.2) of GDP 001 Induction Procedure. 8. Attachments: 8.1 The University of Newcastle PC2/PC3 Facilities 8.2 LSB Safety Notice 9. Check List: 9.1 Necessary records have been signed. 10. References: 10.1 Australian/New Zealand Standard - Safety in laboratories, Part 3: Microbiology. (AS/NZS : 1995) See the Resource Manager CCH Laboratory Safety Manual (LS3-26) CRC Handbook of Laboratory Safety, 3rd Edition. (LS3-26) The University of Newcastle Occupational Health and Safety Committee - Guidelines for the Safe use and Disposal of Biological Material and Material of Biological Origin. (Stored in the Medical Biochemistry Reference Folder, LS3-26). 11. Change History: 11.1 Issue Number: 1st Issue Date Issued: 4 th October Issue Number: 2nd Issue Date Issued: 1st March 2005 Reason for change: Change of roles within Faculty, change to General procedure for School of Biomedical Sciences