Introduction

The United States Department of Agriculture (USDA) and the Public Health Service (PHS) require that survival surgery performed on animals be conducted using aseptic technique.  The requirements of the USDA and the PHS (as defined by the Association for Assessment and Accreditation of Laboratory Animal Care – AAALAC) for survival surgery in rodents and other small animals are summarized below in the “Requirements for Small Animal Survival Surgery”.  Survival surgery is defined as any surgical procedure from which the animal will recover, even if only for short periods of time.

These requirements apply to survival surgical procedures in small mammals belonging to the Order Rodentia and the specific Chiroptera (bat) species, Pteronotus parnelli, Carollia perspicillata and Antrozous pallidus with additional conditions as noted in the document.  The conditions are also applicable to survival surgery in small amphibians, reptiles, and fish with appropriate modifications.  Other species may be added as needed with IACUC approval.

The IACUC recognized that unusual circumstances may necessitate the development of alternative approaches to address specific research needs.  However, IACUC approval is necessary before implementation of variations from the “Requirements”.

As with any activity involving animals the individual carrying out the procedure must be adequately trained.  In some cases it may be necessary for the surgeon to practice the technique on cadavers before performing it in a living animal.  Please feel free to contact the Comparative Medicine Unit at X-6555 or at (330) 325-6555 to arrange for guidance and practice.

In the list below explanatory information is provided in italics for several requirements.  Recommendations intended to assist in the implementation of this policy and enhance the well being of the surgical patient follow the requirements.

Requirements for Small Animal Survival Surgery

1.     The surgery area must be dedicated to that purpose while surgery is conducted. 
Surgery on bats must be performed in an area dedicated solely for that purpose.

2.    The surgery table and associated equipment (e.g., stereotaxic apparatus) must be sanitized prior to use.

3.      Surgical instruments and implantable materials must be sterilized by an approved method.  Instruments used in bat surgery must be autoclaved.

Sterilization may be achieved by autoclaving (minimum 121 °C, 15 PSI, for 15 minutes), dry heat (171°C for one hour), ethylene oxide, irradiation, chemical sterilization according to the product manufacturer’s recommendations (e.g., Cidex^®, or 10% providine iodine) or other means as approved by the IACUC.  Autoclaving generally is the preferred method for sterilization because of its convenience, low cost, and efficacy.  Depending upon the nature of the surgical procedure, the degree of instrument contamination during the surgery, and the type of animal, a sterile set of instruments may be used for up to 2-5 small animals during the same surgery session.  Between animals the instruments must be decontaminated using a chemical disinfection or a point heat source such a glass bead sterilizer.  Chemical sterilants should be removed from instruments by wiping or rinsing with sterile water or saline to avoid chemical tissue damage.

For larger rodents such as rats or guinea pigs washing with an iodine- or chlorhexidine-based surgical soap (e.g., Betadine® scrub or Nolvasan® scrub) followed by rinsing with clean water and disinfecting with 70% alcohol or iodine solution is acceptable.  For mice and small bats, three applications of 70% alcohol or two of alcohol followed by an iodine solution can be used.  Animals should not be excessively wetted due to the potential for hypothermia.

6.     Volatile anesthetic agents must be suitably scavenged.

7.   Animals must be monitored until they have recovered satisfactorily from anesthesia, i.e., normal respiration, sternal posture and moving about.

8.    The date and a brief description of the surgical procedure, including any drugs administered and the anesthetic agent(s) used, must be noted on the animal’s cage card.

9.     Animals must be monitored post-surgically as often as necessary to assure their well being.  Any abnormal findings must be recorded on the cage card.

Preparation of the Surgical Area

1.    Covering the surgical surface with a clean paper (e.g., plastic-backed lab bench paper) or cloth will help prevent hypothermia and absorb fluids.

2.   The surgery area should be separate from high traffic areas and free of unrelated equipment and supplies.  Laminar flow workbenches such as the Stay-Clean L/F Workbench by Lab Products are useful for small animal survival surgery.

3.   If possible, the surgery area should be subdivided into separate areas for animal preparation, surgery and recovery.

Preparation of the Surgical Instruments and Supplies

1.    The same methods used to sterilize surgical instruments are applicable to implantable materials.  Some materials may be commercially available (e.g., polyethylene tubing) as a sterile product.

2.      Instruments and other autoclavable supplies such as gauze pads and drape material can be easily wrapped with disposable paper wraps designed for that purpose or reusable cloth towels/drapes.  Autoclave confirmation tape should be used on each pack that is autoclaved.  Autoclaved instruments are considered sterile for variable lengths of time depending on the manner in which they are wrapped.  Double cloth wrapped instruments stored in an enclosed cabinet are recognized as sterile for up to 7 weeks unless the integrity of the wrapping is compromised.  Scalpel blades should be purchased sterile and not autoclaved as they are dulled by autoclaving.

3.    Surgical instruments should be cleaned with an instrument cleaner, rinsed, and dried after each surgery session.  A soft toothbrush is often useful for delicate instruments.  Instruments should be stored such that the cutting edges, tips, and points are protected from damage.

Preparation of the Animal

1.      Food and water are not usually withheld from rodents unless there is concern about ingesta within the gastrointestinal tract as may occur for abdominal surgery.

2.       Preparation of the animal is usually best done in an area close to, but separate from, the surgery area.

3.       Plucking or shaving with electric clippers is preferred techniques for removal of hair.  Depilatories and razor shaving should be used carefully due to the potential for dermal irritation.  Loose hair can be removed with a vacuum, tape, or wet gauze.

4.       The depilated area should extend beyond the surgical margins so as to facilitate the maintenance of aseptic technique during surgery, but not so far as to contribute to hypothermia.

5.       Gauze sponges and Q-tips are convenient means to wash, rinse, and disinfect the surgical site.

Surgical Technique

1.     The surgery should be conducted so as to minimize trauma to the tissues and preserve the sterility of the instruments and the surgical field.  It should be completed as quickly as possible without compromising technique; tissues should be handled delicately and, depending on the nature of the surgery, kept moist with sterile saline.  Sutures and staples should not be placed too tightly.  A subcuticular skin suture pattern will often preclude the chewing and removal of sutures by the animal.

2.     Whenever possible the surgery site should be draped with sterile drapes of cloth, paper, surgical gauze, or clear adhesive vinyl to minimize the risk of contaminating the surgery site.  Care should be taken to avoid placing a drape such that the animal cannot be monitored.  Drapes can have the added benefit of keeping the animal warm.

3.     Animals should be kept warm using an external heat source, particularly for procedures of any significant length (i.e., longer than 30 minutes).  An electric or water circulating (preferred) heating pad or an overhead heat source such as a lamp can be used.  Great care must be taken to prevent overheating or burning the animal.  Some heating pads come with a rectal temperature probe that acts as a thermostat to turn the pad on and off.  In all cases the external heat source should be separated from the animal by a towel or other protective barrier.

4.     Also for prolonged procedures, particularly those accompanied by blood loss, warmed fluid therapy should be administered.  The recommended amount is equal to 1-2 cc per 100g body weight per hour of anesthesia plus any blood loss.  Because of the small size of the patients covered by this policy, the intraperitoneal or subcutaneous routes are usually used.

5.     During the surgery the animal’s respiration, tissue color and response to noxious stimuli should be monitored so that corrective action can be taken promptly if necessary.

Recovery from Surgery

1.     Depending upon the nature and duration of the surgery, it may be necessary to provide the post-operative patient with an external heat source during the recovery period.  As described above, steps should be taken to protect the animal from the heat source.  At the very least, animals should be placed in their bedding or provided with other external insulating material (e.g., a towel).  Animals recovering from anesthesia or surgery should not be recovered in a wire bottom cage.

2.     During the recovery period, the animal’s clinical condition should be monitored.  Specific observations should include the body temperature, respiratory pattern, condition of the surgical wound, and strength and rate of heartbeat.

3.     Rodents can often be stimulated to breathe in the case of apnea using gentle chest compression or inflating the lungs with a rubber bulb (from a pipette) applied to their nostrils.  An oxygen rich environment also may be beneficial.  The drug doxapram can be used to stimulate respiration and heart beat when administered orally at a dose of 0.5-1 mg/100g.  Yohimbine (approx. dose 0.1-0.2 mg/100g by intraperitoneal injection) can be used to hasten the recovery in animals anesthetized with anesthetic combinations containing xylazine.

4.     Animals cannot remain in investigator laboratories or other unapproved housing areas for longer than twelve hours without IACUC approval.

Post-Procedural Monitoring

1.    Depending upon the nature of the procedure and the condition of the animal, post-surgical monitoring may range from once daily for one or two days to multiple times per day for extended periods.

2.    Conditions of observation are reviewed by the IACUC at the time of protocol review.  In some cases, such as when the same procedure is conducted on many animals, alternative methods of record keeping (other than on a cage card) can be used.  Please contact the CMU at X-6555 or at (330) 325-6555 to discuss alternative means of record keeping.