Biosafety Containment Level 2

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1 RED DEER COLLEGE, SCIENCE DEPARTMENT Biosafety Containment Level 2 Standard Operating Procedures 10/7/2011

2 Contents HANDLING OF PATHOGENS IN A CONTAINMENT LEVEL 2 LABORATORY INTENT DEFINITIONS ROLES AND RESPONSIBILITIES Faculty members/laboratory Supervisors Waste Generators Staff/Students LABORATORY BIOSAFETY GUIDELINES Working in a Biological Containment Level 2 Laboratory Guidelines of CL2 Laboratories PROCEDURES Personal Protective Equipment (PPE) Working in a Biological Safety Cabinet (BSC) Avoiding the Production of Aerosols Decontamination And Disposal Of Biohazardous Waste Spill Procedures EXPOSURE CONTROL PLAN APPENDICES Application Form for a Biosafety Operating Permit Biosafety Resume for Authorized Workers Inventory Guidelines for Cleaning Equipment or Area Destined for Release Equipment / Area Release Form Culture Tracking Form Autoclave Efficiency Monitoring

3 HANDLING OF PATHOGENS IN A CONTAINMENT LEVEL 2 LABORATORY 1. INTENT This Biosafety Standard Operating Procedure (SOP) applies to all faculty, staff, and students at Red Deer College. The standards and practices listed in this SOP apply to all containment level 2 biological laboratories. It was developed to ensure that work with biohazardous materials is conducted in a safe manner reflecting best practices and adheres to the Laboratory Biosafety Guidelines published by the Public Health Agency of Canada. 2. DEFINITIONS Containment Level 2 (CL2) Laboratory A laboratory that handles Risk Group 2 agents. The primary exposure hazards associated with organisms requiring the CL2 are through the ingestion, inoculation and mucous membrane route. Level 2 agents are not generally transmitted by airborne routes, but care must be taken to avoid the production of aerosols or splashes. Primary containment devices such as Biological Safety Cabinets are to be used, as well as appropriate Person Protective Equipment. All persons entering the facility must be informed of the hazards present. Notifiable Biological Substances: Genetically modified micro-organisms or genetic combinations which are not known to occur naturally Permit Holder: A Principal Investigator or Faculty Member or Laboratory Supervisor who is authorized to work with Biological Substances and/or Biohazardous Materials by the Biosafety Advisory Committee Risk Group 1 Organisms Any biological agent that is unlikely to cause disease in healthy workers or animals. These organisms are referred to as low individual and community risk as they are unlikely to cause disease in healthy workers or animals. Risk Group 2 Organisms Any pathogen that can cause human disease but, under normal circumstances, is unlikely to be a serious hazard to laboratory workers, the community, livestock, or the environment. Laboratory exposures rarely cause infection leading to serious disease; effective treatment and preventive measures are available, and the risk of spread is limited. They are referred to as moderate individual risk, low community risk. * For a list of organisms and their Risk Group go to /page-20.html#h-24 3

4 3. ROLES AND RESPONSIBILITIES 3.1 Faculty members/laboratory Supervisors Faculty members and lab supervisors are responsible for ensuring that: their work areas are compliant at all times with this SOP, RDC Biosafety and Health, Safety and Wellness policies, and applicable federal and provincial health, safety, and environmental regulations. they register and/or obtain the required permits for any activities that use Risk Group 2 substances, biohazardous material, and notifiable substances staff and students have been given adequate safety orientation, site specific training for the work assigned, and instruction on the potential hazards associated with working with biological agents staff and students are competently supervised and have reviewed the department policies and procedures for safety staff and students have access to any hazard information on the substances being used staff and students are provided with the appropriate safety equipment and personal protective equipment necessary to protect their health and safety, and that they know how to use and maintain the equipment biohazardous agents have been disposed of properly 3.2 Waste Generators Waste generators are any persons who generate biohazardous waste as a result of work conducted, including laboratory supervisors, faculty members, technologists, and students. They are responsible for: collecting the waste in accordance with RDC s disposal procedures (see 5.4.3) ensuring that waste is properly segregated, identified, and labelled for disposal keeping an inventory of the waste 3.3 Staff/Students Staff and students working in labs are responsible for ensuring that they: conduct their work in a safe and responsible manner in order to protect their health and safety as well as that of others who may be affected by their acts or negligence are familiar with the potential hazards and this SOP as it relates to working with biohazardous agents promptly report any known accidents/incidents, spills, or unsafe conditions to their supervisor 4

5 4. LABORATORY BIOSAFETY GUIDELINES 4.1 Working in a Biological Containment Level 2 Laboratory The most important element of containment is strict adherence to standard microbiological practices and techniques. Persons working with infectious agents or infected materials shall be aware of potential hazards and shall be trained and proficient in the practices and techniques required for safely handling such material. A CL2 laboratory is suitable for work involving agents of moderate potential hazard to personnel and the environment. Working in a CL2 laboratory involves: working under the supervision of personnel that have training in handling pathogenic agents having controlled and limited access to the laboratory taking extreme precautions with contaminated sharp items special procedures when there is a possibility of infectious aerosols or splashes being created Persons who are at increased risk of acquiring infection, or for whom infection may have serious consequences are not allowed in the laboratory. The laboratory instructor/supervisor has the final responsibility for assessing each circumstance and determining who may enter or work in the laboratory. 4.2 Guidelines of CL2 Laboratories all guidelines of general laboratory safety must be followed (see Biology Laboratory Safety Procedures) all personnel must be trained and understand the hazards appropriate personal protective equipment must be worn personal items (e.g. coats, purses, backpacks) must remain outside the laboratory doors to the laboratory must be closed at all times, and locked when the lab is not occupied hands must be washed after removing gloves and before leaving the lab work surfaces must be disinfected before work begins and after it is completed all mobile equipment must be decontaminated before being removed from the area aerosol production is to be minimized all contaminated or infectious material must be decontaminated before leaving the area use of cell phones is prohibited access to laboratories is limited to authorized personnel only hazard warning signs must be posted efficacy monitoring of autoclaves using biological indicators must be done regularly and records of those results and cycle logs must be kept on file all accidents/incidents, spills, or unsafe conditions must be reported in writing 5

6 leak proof containers are to be used for the transport of infectious materials within the facility in order to release a laboratory from CL2, complete decontamination of work surfaces, fridges, incubators, and equipment must be completed 5. PROCEDURES 5.1 Personal Protective Equipment (PPE) a lab coat, long pants, and shoes that cover the entire foot are to be worn at all times while working in the laboratory gloves must be worn at all times while working with biohazardous materials. Under no circumstance should they be worn outside the lab, and must be disposed of in the appropriate biohazard waste container lab coats must remain in the lab. Under no circumstance should they be worn outside the lab, until they have been properly decontaminated 5.2 Working in a Biological Safety Cabinet (BSC) Biological Safety Cabinets (BSCs) are among the most effective and the most commonly used primary containment devices in laboratories working with infectious agents. They provide protection to the worker as well as to the work being performed. All procedures must be performed carefully to minimize the creation of aerosols. Procedures that create aerosols must be conducted in a Biological Safety Cabinet. Operation of a BSC: Before using the cabinet: if the room is occupied, do not turn on the UV light. If the UV light is used, ensure that it is turned off before continuing. disinfect work surfaces with Oxivir Tb. place required items inside the cabinet turn on the blower and allow to run 5 minutes use of an open flame in the BSC should be kept to a minimum. After completion of work: allow the blower to run for 5 minutes in order to purge the BSC remove and decontaminate materials and equipment disinfect work surfaces turn off the blower and fluorescent light, and turn on the UV light if room is not occupied For more detailed instructions on using a BSC, refer to Operation and Maintenance Manual (located in drawer ). 6

7 5.3 Avoiding the Production of Aerosols Centrifuge Operation If a microorganism is not contained within a centrifuge, aerosols can be produced during the centrifugation process. This could occur with uncapped samples, or when a leak, spill or breakage of the tube occurs. Minimize the risks when centrifuging by following the guidelines below: use unbreakable tubes avoid overfilling the tubes use caps or stoppers on centrifuge tubes use outer, sealable safety cups that can be loaded and unloaded in a BSC decontaminate the outside of the cups or buckets before and after use and inspect seals regularly for deterioration ensure that the centrifuge is properly balanced do not open the lid during or immediately after operation allow the centrifuge to come to a complete stop before opening Mixing Operations Sonicators, shakers and homogenizers can generate aerosols during operation. Minimize the risks when mixing by following the guidelines below: use sealed vessels during mixing and allow the vessels to settle before opening open mixing vessels inside a BSC check the condition of the mixing equipment routinely for deterioration disinfect all exposed surfaces before and after use Vacuum and Aspirating Equipment Minimize the risk of generating aerosols during vacuum and aspiration operations by following the guidelines below: use non-breakable equipment (i.e. do not use glass) ensure that vacuum equipment is fitted with a HEPA filter place a disinfectant in the overflow flask of the aspirating equipment Needles and Syringes Minimize risk while using needles and syringes by following the guidelines below: perform all operations with needles and syringes in a BSC fill syringes carefully to avoid frothing or introducing air bubbles use blunt-end needles for removal or introduction of fluids through small apertures in equipment dispose of needles and syringes into appropriate sharps container Pipettes Minimize risk while using pipettes by following the guidelines below: use cotton-plugged pipettes 7

8 keep pipettes upright while in use and between steps to prevent contamination of the mechanical suction device and work surface gently expel the contents of the pipette close to the surface of the liquid to allow to flow down the side of the container do not mix contents of a container by alternating suction and expulsion with the pipette to deliver (TD) pipettes are recommended vs. to contain (TC) pipettes so the last drops do not need to be expelled for an accurate measurement TC pipettes are to be used in the BSC due to the potential for aerosol production submerge used non-disposable pipettes horizontally in disinfectant solution to avoid forcing out the any liquid remaining in the pipette Transfer Loop Minimize risk while using transfer loops by following the guidelines below: when disinfecting the loop between inoculations, hold the loop close to, but not in the flame until the material has charred and then fully insert into the flame substitute an enclosed micro-incinerator for an open flame burner or disposable loops where possible 5.4 Decontamination And Disposal Of Biohazardous Waste Chemical Disinfection Chemical disinfectants are used for the decontamination of surfaces and equipment that cannot be autoclaved, any item for which heat treatment is not feasible, and for cleanup of spills of infectious materials. Oxivir Tb may be used for disinfection Autoclave Infectious laboratory wastes (Petri dishes, pipettes, culture tubes, glassware) can be effectively decontaminated in an autoclave. Operation of an Autoclave turn the autoclave on and allow to warm up for ½ hour place items in the chamber. Bags must be unsealed and must be placed in leak proof autoclavable trays place an indicator strip in appropriate area of the autoclave chamber close door choose the appropriate cycle depending on the items and the size of the load press start when the cycle is finished, open the door carefully to avoid escaping steam wearing heat resistant gloves, remove items and place decontaminated labels on any items to be placed for general garbage pick-up Efficiency testing must be performed on the autoclave on an on-going basis. See the Autoclave Efficiency Monitoring form in the appendix. 8

9 For more detailed information on the operation of the autoclave refer to Operator Manual (located in instructors desk, room 1430) Disposal All biohazardous wastes must be disposed of in the appropriate labelled containers. contaminated sharps (needles and syringes, broken glass, microscope slides) must be placed in rigid, puncture resistant, leak-proof, closable sharps containers labelled with WHMIS approved biohazard symbol contaminated disposable non-sharps (Petri plates, culture tubes, gloves, paper towels) must be placed in bags labelled with WHMIS approved biohazard symbol contaminated non-disposable items (instruments, glassware) must be placed into appropriately labelled containers After proper disposal, contaminated items must be autoclaved or otherwise decontaminated in a timely manner. 5.5 Spill Procedures Inside a BSC remove gloves inside the BSC and leave the area for at least 5 minutes wear lab coat, safety glasses, and gloves during clean-up place contaminated reusable items in autoclave bags or trays cover spill with absorbent paper towels and soak with disinfectant (Oxivir Tb). Allow at least 10 minutes contact time place towels and any other disposable items in autoclave bag wipe down work surfaces, walls, and equipment in the BSC with disinfectant (Oxivir Tb) allow the BSC to run 10 minutes before resuming work or turning it off Outside a BSC clear area of all personnel, and wait at least 30 minutes for the aerosol to settle remove contaminated clothing and shoes and place in a biohazard bag put on a lab coat, safety glasses, and gloves cover spill with absorbent paper towels and soak with disinfectant (Oxivir Tb). Allow at least 10 minutes contact time decontaminate all items within the spill area place paper towels and any other disposable items in an autoclave bag Reporting all spills must be reported on an Incident/Injury Report Form. This form can be found on the Red Deer College website, Log in to the Loop, click on Quick Access and then Forms Index. 9

10 6. EXPOSURE CONTROL PLAN The Permit Holder shall develop a written Exposure Control Plan (ECP) for situations where any student or worker is required to handle, use, or produce hazardous or infectious materials. This plan shall: identify any student or worker at the place of employment who could be exposed to hazards in the workplace describe the ways in which material can enter the body and the risks associated with that entry describe the signs and symptoms of any illness that may arise from exposure describe (referencing this SOP) the safety control measures and their limitations, including: o administrative safety procedures o procedural safety controls and training o engineering safety controls and training o PPE (include training & maintenance) o immunization in accordance with the University Immunization Policy and o the use of Universal Precautions when handling blood and body fluids. describe (referencing this SOP) emergency response procedures for: o spills or leaks of hazardous materials o when a worker has been or believes they have been exposed to a hazardous material describe (referencing this SOP) methods of cleaning, disinfecting, or disposing of clothing, PPE or other equipment if contaminated with hazardous materials, and indicate who is responsible for carrying out those activities describe the training that will be provided to students or workers and the means by which this training will be provided require the investigation and documentation of any work-related exposure incident, including the route of exposure and the circumstances under which the exposure occurred require the investigation of any occurrence of an occupationally acquired illness or an occupationally transmitted infectious disease in order to identify the route of exposure and to implement measures to prevent further illnesses 10

11 APPENDICES 11

12 Application Form for a Biosafety Operating Permit Name of Applicant: Position: Department: Office Phone #: Project Description: Attach a brief summary of the experimental procedures including an inventory of the names of any biohazardous materials or notifiable biological substances used indicating their Risk Group Level. Identify what species the agent could infect, comment on its virulence, hosts, vectors, routes of infection, etc. State what potential impact the biohazardous materials or notifiable biological substances may have on the health and safety of personnel, the public and the environment. Develop a written Exposure Control Plan for situations where a student or worker is required to handle, use, or produce biohazardous or infectious materials. ( ) Check this box to confirm that you will be working in an appropriate containment facility and using the appropriate microbiological procedures as set forth in the "Laboratory Biosafety Guidelines" (Health Canada). If NOT applicable, check ( ). Attach any permit (import/export) restrictions or conditions associated with this work. Work Location and Containment Level: Room #s: Outdoor locations identified if applicable: Agent Storage Locations (i.e. freezers etc): Containment Level Assigned to area: Level 1 ( ) Level 2 ( ) Level 3 ( ) Level 4 ( ) Do you have a BSC: Yes ( ) No ( ) Other Containment Required: Yes ( ) No ( ) Cabinet Model Number: Serial Number: Date Last Tested: Is there controlled access to the work area?: Yes ( ) No ( ) Authorized Workers: Attach a resume form for each worker and permit holder. I accept responsibility for the accuracy of the information in this application and ensure that this project will be performed in accordance with the RDC Biosafety Standard Operating Procedures. Signature of Applicant: Approval of the Biosafety Officer: 12 Date: Date:

13 Biosafety Resume for Authorized Workers (PLEASE PRINT OR TYPE ALL OF THE FOLLOWING INFORMATION) Personal Information: Name: Phone: Education and Experience: Department: Room: Degree(s), Diploma(s) & Position: Number of years worked with biohazardous materials or notifiable biological substances: Training: List any formal Biosafety courses taken (Date, Length, Location): Please verify the following: have attended the RDC biosafety course principles and practices of biological exposure/infection controls are followed have read the Laboratory Biosafety Guidelines (Health Canada) or watched the Lab Biosafety 101 DVD know the biological effects of exposure (signs & symptoms) and the ways it can occur information (or PSDS) is available on the biohazardous materials or notifiable biological substances used understand the function/use and maintenance of biosafety cabinets and personal protective equipment (PPE) understand the emergency procedures that are in place for exposure, accidents, incidents, and spills with a written recording and follow up system will not remove Risk Group 2 organisms from the Containment Level 2 labs have read the Red Deer College Biosafety Policy and the SOP on Level 2 Containment understand the Right to Know the hazards in the workplace under the Health and Safety Legislation understand the Right to Refuse unsafe acts under the Health and Safety Legislation I have read the Biosafety Standard Operating Procedures and understand and accept my duties and responsibilities in accordance with the Biosafety Standard Operating Procedures. Signature of Authorized Worker: Date: 13

14 Inventory Biological Material or Agent Host Ranges Containment/ Risk Group Level 2. 14

15 Guidelines for Cleaning Equipment or Area Destined for Release Objective The objective of the Equipment or Area Release guidelines are to protect the health and safety of all staff, students, and the public at large from being exposed unnecessarily to equipment or areas that may contain hazardous biological substances. It is hoped that the person requesting service in an area or on a piece of equipment will take the time to evaluate the associated risks from the point of view of the service technician who will be completing the work. Biological Substances The minimum for cleaning any area or equipment contaminated with biological agents is for the technical staff in the laboratory to wipe down the area or equipment with a disinfectant that is effective on that biological substance (Oxivir TB). Equipment shall be autoclaved if this is a feasible option. Personal Protective Equipment and Precautions Personal protective equipment and precautions shall be taken in handling equipment or upon entering a certain area. Ask technical staff what you should wear when handling or entering the designated area to work. Always protect your eyes and hands. Wash your hands and tools upon completion of the work. Consultation The Office of Health, Safety and Wellness will provide consultation if there is any unresolved safety concerns. Examples Equipment that may require decontamination and the use of personal protective equipment are: biosafety cabinets vortex mixers fume hoods sonicators autoclaves shaker units incubators centrifuges refrigerators pipette pumps water baths lab coats spectrophotometers Areas that may require decontamination and the use of personal protective equipment are: laboratories prep rooms fume hood /BSC discharge areas 15

16 Equipment / Area Release Form Supervisors, Technologists, and Faculty members releasing equipment or areas for service work, relocation, disposal, or resale which may contain biohazardous residue within or on the surface are required to complete an Equipment /Area Release Form. This form may be attached to the equipment or given to the service employee. Room #: Description of Area /Equipment: Model #: Serial #: (attach list if necessary) Destination: ( ) Service ( ) Relocation ( ) Disposal ( ) Area turnover Comments: I declare that the equipment /area specified above has been cleaned and decontaminated so that it does not present any hazard associated with biohazardous substances. Signature: Date: Phone #: 16

17 Culture Tracking Form Culture: Date Acquired: Subculture Date Batch # Purpose Initials Disposal Date Initials 17

18 Autoclave Efficiency Monitoring Date Bowie-Dick Test (SMART Pack) Biological (Biosign Biological Indicators) Date Bowie-Dick Test (SMART Pack) Biological (Biosign Biological Indicators) Daily: Clean sediment screens; blow down boiler; Bowie-Dick Test If Bowie Dick Test fails, perform Vacuum Leak Test Weekly: Biological Control testing 18

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