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STANDARD OPERATING PROCEDURES

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1. Performance Standard: Practices and procedures used when working with animals at the ABSL2 level are in accordance with those described in this SOP and the CDC-NIH Biosafety in Microbiological and Biomedical Laboratories, 4th Edition.

2. Standard Practices

• All personnel working at the ABSL2 hazard level are required to attend the College's BSL2 training program. The Animal Resources Center (ARC) staff attends an ABSL2 training. The ARC and Environmental Health & Safety (EH&S) will provide additional training as needed. The CDC/NIH Biosafety in Microbiological and Biomedical Laboratories, 4th Edition, is used as the basis for education and training. This document is available on the EH&S web site at: www.dartmouth.edu/~ehs/
• Eating, drinking, smoking, handling contact lenses, applying cosmetics, and storing food for human use are not permitted in the animal facilities.
• Access to the animal facility is limited or restricted to personnel who have been advised of the potential hazard and who need to enter the room for program or service purposes when work is in progress. Access is at the discretion of the Animal Care and Use Program (ACUP) Director or Associate Director. The PI is responsible for advising his/her personnel on the potential risks due to the hazards involved in the project. The ARC is responsible for advising its personnel on the potential risks due to the hazards involved in the project. Persons who may be at increased risk of acquiring infection, or for whom infection might be unusually hazardous, are not allowed in the animal room.
• All personnel must receive appropriate immunizations or tests for the agent handled or potentially present (e.g., hepatitis B vaccine or TB skin testing). Occupational Medicine at Dartmouth Hitchcock Medical Center (DHMC) will be consulted as appropriate, depending on the agent under study and the potential risk to personnel.
• When appropriate, considering the agent handled, baseline serum samples from animal care and other at risk personnel are collected and stored. Additional serum samples may be collected periodically depending on the agent handled or the function of the facility.
• A study requiring ABSL2 practices is performed in an animal room dedicated for this purpose after discussion between the Principal Investigator (PI) and the ACUP Director or Associate Director.
• Research personnel are required to
  • be listed on an IACUC approved protocol,
  • have completed a Personnel Information Form,
  • be enrolled at the appropriate level in the ARC Occupational Health Program,
  • have attended the ACUP Orientation and tour for access into the animal facility and
  • have received individual training from the ARC staff for entering an ABSL2 room.
• An IACUC and EH&S approved protocol describing the procedures and a Special Needs Summary Sheet (SNSS) for the study are required prior to ordering animals on any protocol involving hazards.
• The ARC and EH&S training must be completed and the lab must be in compliance prior to the start of the ABSL2 work.
• The PI/research staff is responsible for putting a biohazard sticker, including the name of the biohazard, on the cage card.
• The ARC's Operations Manager is responsible for
  • posting a hazard warning sign incorporating the universal biohazard symbol on the access door to the animal room when an infectious agent is in use in the animal room. The hazard warning sign identifies the infectious agent(s) in use; lists the name and telephone number of the responsible animal facility personnel, the PI, and other responsible person(s); and indicates the special requirement(s) for entering the animal room.
  • posting an approved copy of the SNSS on the animal room door.
• Use caution when handling contaminated sharp items (e.g. needles and syringes, slides, pipettes, capillary tubes, and scalpels). Substitute plastic ware for glassware whenever possible.
  • Use only needle locking syringes or disposable units where the needle is integral to the syringe.
  • Syringes that re-sheathe the needle, needleless systems, and other safe devices should be used when appropriate. Training for use of these systems is provided by the ARC veterinary staff or EH&S as necessary.
  • Do not be bend, shear, break, recap, remove or otherwise manipulate by hand needles from disposable syringes. Carefully place them in the sharps container provided by the ARC staff.
  • Place non disposable sharps into a hard walled container for transport to a processing area for decontamination.
  • Do not handle broken glassware directly by hand. Removed it by mechanical means such as a brush and dustpan, tongs, or forceps.
  • Containers of contaminated needles, sharp equipment, and broken glass should be decontaminated before disposal, according to the EH&S "Green Guide".
• Place cultures, tissues, or body fluid specimens in a container that prevents leakage during collection, handling, processing, storage, transport, or shipping.
• Contaminated equipment must be decontaminated according to local, state, or federal regulations before it is sent for repair or maintenance or packaged for transport in accordance with applicable local, state or federal regulations before removal from the facility.
• Decontaminate cages and accessories before they are brought to Cagewash. Autoclaving is the preferred method of decontamination. If autoclaving is used, perform biological monitoring on a weekly basis to verify cycle parameters. If autoclaving is not practical or possible, thoroughly decontaminate cages and accessories with Wescodyne (for rodents) or Virkon S (for non-human primates) following the mixing ratio given below.
  • Wescodyne is mixed at a rate of 3 ounces Wescodyne to 5 gallons of warm water. A minimum of 10-minutes contact time is required.
  • Virkon S is mixed at a rate of 2 packages to 1 gallon of water. A minimum of 10-minutes contact time is required. Note: Virkon S must be made fresh each day.
• Equipment and work surfaces should also be decontaminated using paper towels saturated with Wescodyne (for rodents) or Virkon S (for non-human primates), after work with infectious materials is finished and especially after overt spills, splashes, or other contamination by infectious materials. Wear nitrile gloves when using Wescodyne and/or Virkon S. Dispose of the gloves and paper towels in the biohazard trash.
• Perform all procedures carefully, to minimize the creation of aerosols. Aerosol-producing situations include routine husbandry procedures and inoculation of animals with infectious organisms. Regarding husbandry procedures, the following precautions should be taken to minimize aerosol production:
  • Floors are cleaned by application of bleach solution to the floor (1/2 cup of bleach in 1 gallon of water), brooming it around to cover the floor, and hosing the floor to remove the bleach solution. Pour 1 cup of bleach in the floor drain trap after the room floor is rinsed.
  • Bedding from animals requiring ABSL2 procedures is dumped into a black bag using a biosafety cabinet. The bag must be sealed, labeled with an EH&S Associated/Additional Hazard card, and placed in the infectious waste box for disposal.
• Aerosol-producing activities, including necropsy of infected animals, harvesting of tissues or fluids from infected animals or eggs, intranasal inoculation of animals, and manipulations of high concentrations or large volumes of infectious materials, must be done in a biological safety cabinet unless special precautions have been approved in advance.
• Double-bag all wastes that can be effectively incinerated and attach an EH&S Associated/Additional Hazard card to the bag. Spray the outside bag with Wescodyne (for rodents) or Virkon S (for non-human primates) prior to removing it from the room.
  • Place bags that don't require refrigeration/freezing in an infectious waste box.
  • Places bagged carcasses in the cooler/freezer labeled as a storage site for biohazard samples.
• Personnel must remove their gloves and thoroughly wash their hands after handling cultures and animals, and before leaving the animal facility. Hands must be washed with an antimicrobial soap for at least 30 seconds, in the nearest sink, prior to leaving the ARC. The ARC provides a hand sanitizer when a sink is not available in the animal facility.
• An insect and rodent control program is in effect. The routine Animal Resource Center pest management program applies to all areas where ABSL 2 studies may be performed.

3. Special Practices

• The study will not begin until training specific to the study is completed and documented by the ARC. Personnel receive annual updates, or additional training as necessary for procedural or policy changes. The IACUC Manager maintains records of all training, as per CFR 1910.1030.
• Animals not involved in the biohazardous study are not permitted in the animal room.
• All equipment leaving the room is appropriately decontaminated as described in section 2 of this SOP, above.
• When needed, animals are housed in primary biosafety containment equipment appropriate for the species.
• Spills and accidents that result in overt exposures to infectious materials must be immediately reported to the ARC, DHMC Occupational Medicine, and to the PI. Medical evaluation, surveillance, and treatment are provided as appropriate and written records are maintained.

4. Animal Facility

• The animal facility is separated from areas that are open to unrestricted personnel traffic within the building. Access to the facility is limited by a key-card access security system.
• The ARC facilities are designed and constructed to facilitate cleaning and housekeeping.
• A handwashing sink is available within the animal facility where infected animals are housed. However, because the sink may not be in the same area as the biohazard, the ARC also provides a hand sanitizer.
• Exhaust air is discharged to the outside without being re-circulated to other rooms. The direction of airflow in the animal room is inward.
• An autoclave that can be used for decontaminating infectious waste is available within the Borwell and Vail animal facilities.

5. Standard Personal Protective Equipment (PPE)

• See section 6 of this SOP for details of the various risk levels and PPE required when working with non-human primates.
• All personnel entering animal rooms housing ABSL2 agents must wear the following PPE:
  • disposable gown,
  • nitrile gloves (double gloves are worn when there is a potential of scratch hazard),
  • shoe covers (two pairs),
  • bouffant cap and a
  • face mask with face/eye (mucous membrane) protection.
  • NOTE: PPE is put on outside of the animal room and removed immediately prior to exiting the animal room or in the anteroom. The outer pair of shoe covers must be removed as you are stepping out the door while leaving the animal room. With the exception of the inner pair of shoe covers, PPE is disposed of as infectious waste. Cut resistant gloves (Kevlar) can be worn over disposable gloves, if appropriate, and decontaminated and sanitized as necessary.

6. Non-Human Primate Personal Protective Equipment

• PPE will be used by personnel that enter the non-human primate housing room, and by personnel working in the laboratory with the non-human primates. The following are minimal recommended requirements for PPE based on the relative risk:
  
  High Risk--This would include any activity that involves high possibility of a splash hazard (i.e. hosing down cages, hosing the room, removing waste pans, etc.).
  Wear the following:
  Work clothes (scrubs)
  Lab coat with stretch cuffs, designated for primate use.
  Nitrile gloves (double)
  Shoe covers (waterproof) or rubber boots
  Non-disposable face shield and safety glasses or goggles
  Face mask (fluid repellant) covering nose and mouth
  Bouffant cap
  
  Moderate Risk--This would include activities that do not involve a splash hazard, but more than simple observation of the animals with no contact (removing the primate from the cage for studies, replacing the primates back into the cage, cylinder cleaning, physical exams on an anesthetized animal, etc.)
  Wear the following:
  Lab coat with stretch cuffs (over street clothes), designated for primate use.
  Nitrile gloves
  Shoe covers (waterproof)
  Safety glasses or goggles
  Face mask (fluid repellant) covering nose and mouth
  Bouffant cap
  
  Low Risk--This would include animal observation in the laboratory while the primate is completely contained in the primate perch.
  Wear the following:
  Lab coat with stretch cuffs (over street clothes), designated for primate use.
  Nitrile gloves (single)
  Safety goggles or safety glasses
  Face mask (fluid repellant) covering nose and mouth
  Bouffant cap
  
  
• Dedicated non-human primate lab coats are disinfected through the normal laundry process.

7. Use transfer cages to transfer non-human primates between cages for cage changes. Kevlar lined non-human primate gloves and a net are available in the anteroom and used if necessary to help capture escaped non-human primates.

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