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Principles of Asepsis

Preparation- General

• Instrument Prep
• Patient Prep
• Surgeon Prep

Surgical Technique

Post-procedural Care

Antibiotic Guidelines

Sample Rodent Surgery SOP

CDC Guidelines for Prevention of Intravascular Device-Related Infections 

CDC Guideline for Prevention of Surgical Site Infection
 

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Experimental Surgery

General Issues and Requirements

Surgery is classified in several ways.  There are different requirements depending on the type of surgery being performed.

• Surgery is major if it  enters a body cavity (thorax, abdomen, calvarium), or has the potential for having significant complications.  Included would be orthopedic procedures and extensive cannulation procedures.
• Other surgery is classified as minor. Minor procedures include peripheral vessel cannulations and skin incisions.
• Surgery is also classified as survival vs. nonsurvival.  Asepsis and sterility are not required for non-survival procedures, unless the procedures are of sufficient duration to allow bacterial infections to affect the outcome of the study.
• There are also slightly different requirements for surgery performed on large animals such as rabbits, dogs, pigs and monkeys versus rodents and non-mammals.

Surgery Facilities

• Separation of the preparation areas from the surgery area
• Minimization of personnel traffic flow through the surgery area
• Air flow should be away from the surgery area (e.g. positive room pressure, use of filtered, laminar flow air).  It may be desirable to have HEPA filtered air for high-risk procedures
• Room surfaces should be non-porous and easily sanitized
• A regular room cleaning and disinfection schedule should be established (i.e. daily cleaning of floors and work surfaces, weekly to monthly cleaning of walls and cabinets)
• The surgery area should be free of all equipment and materials not necessary for the procedure.  Any stored items should be in cabinets or drawers.

• An approved surgery suite is needed for large animal major survival surgery, with separate rooms for preparation of the patient, preparation of the surgeon, the operating room and a recovery of the animal from anesthesia.
• Non-survival surgery, minor surgery or rodent/non-mammal surgery may be performed in a dedicated work area.  This is a room or bench top which from which all materials are removed at the time of the surgery.  The same concepts described above are important for a dedicated area.

Principles of Asepsis

• Time of surgical procedure is an important factor, as the longer a procedure takes the greater the possibility of contamination and therefore infection.
• Trauma that is sustained by the tissue as a result of rough handling, drying out upon exposure to room air, excessive dead space, implants or foreign bodies or non-optimal temperatures will contribute to infections.
• Trash refers to contamination by bacteria or foreign matter.

Preparation: Surgeon, Patient, Instruments and Supplies

• It is essential that anything that will contact the subcutaneous tissues of an animal be appropriately sterilized to prevent post-procedureal infections.  These will be discussed below.  Videos on many of these techniques are available from the central animal facilities and the University library system.  The RAR veterinary staff can provide training as well. Other aspects of preparation include pre-operative fasting, if necessary, a decision about prophylactic antibiotics, appropriate anesthesia of the patient, and a plan for post-operative pain control and supportive care. 

Preparation of Instruments and Supplies

Surgical instruments and supplies must be sterilized before they are used for survival surgery.  There are a number of ways that this can be achieved.
 

• Durable instruments and supplies may be autoclaved.  This an extremely reliable and cost-effective method for sterilization.  The disadvantage is the time that it takes to perform (from 15 minutes to 1 hour). Normally a wrapped "pack" of instruments is prepared and is opened the day of surgery. Packs may be stored if they are kept away from moisture. A preparation date should be put on each prepared pack and packs should not be used if they are more than six months old.
• Instruments and less durable supplies may be sterilized by ethylene oxide.  This is also a reliable method for sterilization.  However, it is more costly than autoclaving and also takes time to perform (overnight).  Ethylene oxide is hazardous and must be performed using appropriate procedures and equipment.  The Fairview-University hospital Fairview Sterile Stores service can provide this service (672-4234 or 672-7181).
• Instruments and some materials may be sterilized in a cold sterilant solution. There are several acceptable commercial sterilants available. Only products classified as sterilants are to be used for sterilizing instruments and implants for surgery and they must be used according to the manufacturer's recommendations for sterilization. Following are examples of four commercial products listed by brand names: Cidex ­ active ingredient: 2% glutaraldehyde; Sporicidin ­ active ingredients: phenol 7.05%, glutaraldehyde 2%, Sodium phenate 1.2%; Alcide ­ active ingredient: sodium hypochlorite; and Sporclenz - for a minimum of 6 hours. active ingredient: hydrogen peroxide. Non-commercial solutions that are acceptable include: glutaraldehyde 2% for a minimum of 10 hours; 8% formaldehyde + 70% ethyl alcohol for 18 hours. All surfaces, both interior and exterior, must be exposed to the sterilant. Tubing must be completely filled and the materials to be sterilized must be clean and arranged in the sterilant to assure total immersion. The items being sterilized must be exposed to the sterilant for the prescribed period of time. The sterilant solution must be clean and fresh. Most sterilants come in solutions consisting of two parts that when added together form what is referred to as an "activated" solution. The shelf life of activated solutions is indicated on the instructions for commercial products. Rinsing chemically sterilized items. Instruments, implants, and tubing (both inside and out) should be rinsed with sterile saline or sterile water prior to use to avoid tissue damage. Note: chemicals classified only as disinfectants (for example, 70% alcohol) are not adequate.
• If starting with sterilized instruments, the same instruments can be used for multiple animals (rodent and non-mammal only) if the instruments are re-sterilized between animals, for example with a hot bead sterilizer. However, because only the tips of the instruments are sterilized, the surgeon must constantly be aware of instrument handling and aseptic technique. The pack of instruments must be fully re-sterilized (autoclaved) if the sterile field is broken, e.g. instruments are set down on a table or other non-sterile surface between surgeries. The efficacy of the hot bead sterilization is high and it sterilizes in a very short time (10 sec) but it is necessary to allow the instruments to cool before handling tissue to prevent thermal injury. [Ref: Callahan, et. al, 1995. A comparison of four methods for sterilizing surgical instruments for rodent surgery. Contemp. Top. Lab. Anim. Sci, 34:2, 57-60.]
• Instruments and materials are often available pre-sterilized.  The packages should have an expiration date on them.  Surgical supplies may not be used for survival surgery when they have passed the expiration date.


Use of Expired Materials

Expired medical materials such as drugs, fluids and sutures may not be used on any research animal who is unanesthetized or who is to recover from an anesthestic procedure. The use of such materials under these conditions constitutes inadequate veterinary care under the Animal Welfare Act. 

The IACUC has established the following guidelines for the use of expired medical materials: 
 

1. It is never acceptable to use outdated anesthetics, analgesics, or emergency drugs. Examples of acceptable materials include IV fluid solutions, non-emergency drugs (diuretics, contrast material, antibiotics), IV catheters, bandage materials, surgery gloves and suture materials. 
2. Expired materials are only to be used on anesthetized animals in terminal studies (e.g. studies from which the animal does no awaken).  Anesthesia for these terminal studies must be induced and maintained using current, non-expired drugs. 
3. All expired materials must be clearly and individually labeled as “Expired--for acute use only” and are kept together in an area physically separate from all other medical materials and drugs. The area (box, shelf etc.) they are kept in must be labeled “Expired--for acute use only”. 

Preparation of the Patient

The majority of post-procedural infections are the result of contamination of the surgical site with resident or transient skin bacteria from the patient.  Therefore, decontamination of the surgical site and prevention of contamination from other areas is the best means of preventing post-procedural infections.
 

• Normally, the patient's hair should be removed from the surgical site.  This should done with an electric clipper or depilatory rather than a razor.  Hair removal should be performed immediately prior to the surgery.  Extended time between hair removal and use of razors contributes to post-procedural infections. 
• The patient's skin should be scrubbed with a disinfectant such as povidone iodine or chlorhexidine. Scrubbing should start at the center of the surgical site and move to the outside in a linear or circular manner. Scrub the surgical site with a disinfectant and rinse/scrub with alcohol or sterile water or saline to remove debris. Repeat at least three times or until site is free of visible debris. Often a disinfectant solution is then painted onto the surgical site and left to dry. The amount of scrub fluid used should be carefully controlled to prevent hypothermia. It may not be appropriate to scrub the site of some patients. Scrubbing the skin of a fish or amphibian will remove the protective bacterial slime layer, and may actually increase the risk of infection.
• A sterile surgical drape should be used whenever possible to isolate the disinfected area from surrounding areas.  To be effective, a drape must fit tightly to the skin and must be impermeable to moisture.  Clamps or sutures may be used to fix the drape in place.  Self-adhesive drapes are also useful and are particularly recommended for use in small patients.  In some cases a drape may not be practical or necessary.  When a drape is not used is places extra responsibility on the surgeon to perform excellent surgical technique.

Preparation of the Surgeon

The patient must be protected from organisms that can be carried and shed by the surgeon.  These organisms reside on the surgeon's skin, hair, in the nose or mouth, or may be carried on dust particles from the floor or room surfaces.  This route of contamination is minor compared to the patient's own flora, however, it is a significant source of contamination is some types of surgery such as orthopedic and central nervous system procedures.
 

• Sterile gloves should be used for all procedures.  Examination gloves are not sterile.  Gloves may be disinfected between surgeries with a cold sterilant for rodent and non-mammal surgeries.  Large animal surgeries should be performed with a new pair of gloves for each patient.
• The surgeon's hands and arms should be scrubbed for 3 minutes with a disinfectant such as povidone iodine or chlorhexidine, rinsed with water and dried prior to gloving for any large animal survival surgery.  As much as 30% of the time gloves become perforated during surgery, exposing the animal's tissues directly to the surgeon's skin.
• A cap, face mask, shoe covers and sterile gown must be worn for all large animal major survival surgeries.
• A clean smock or lab coat is recommended when performing rodent surgeries.  A hair cover and face mask will reduce the risk of gross contamination of the surgical site.
• Minimizing traffic flow and conversation in the operating room significantly reduces the risk of contamination of the surgical site.

Surgical Technique

It has been recognized that one of the greatest influences on the incidence of post-procedural infection rates is the surgeon themselves.  Prolonged surgical times expose tissues to contaminants, dry out tissues and compromise the blood flow to tissues.  Tissues damaged by crushing or drying, suture and other surgical implants serve as a nidus for infection.  There are a number of things that surgeon's can do to prevent post-procedural infections.
 
• Be aware of instrument and hand position at all times.  If an instrument or hand touches something outside of the sterile field (the are delimited by the drape or the inside of the opened instrument pack) the instrument or glove should be replaced immediately.
• Be gentle when handling tissues. 
• Do not use toothed or crushing instruments if it is not necessary.
• Hold the cut edge rather than grasping in the middle of a tissue layer.
• When tying off vessels include only a minimum of surrounding tissues.
• Use electrocautery or electroscalpels sparingly.  They cause significant tissue necrosis.
• Use appropriate suture techniques 
• Any suture that will be buried in tissues should be either absorbable or monofilament (non-absorbable braided suture is irritating and can harbor bacteria)
• Sutures should be placed evenly and as close to the tissue edge as possible to prevent obstruction of blood flow- typically no more than 1 cm from the edge is necessary in large animals and 0.2 cm in small animals.
• Sutures should only be tightened enough to appose the tissue edges.  Any tighter will obstruct blood supply, retard wound healing and may result in dehiscence.
• Skin sutures are often unnecessary.  They may cause the animal to chew or scratch at the incision site.  Alternatives include use of subcutaneous/intradermal closure techniques or tissue adhesive.
• Ablate all "dead space" during closure.  Any pockets or potential space that remains between tissue layers will fill with extracellular fluid or blood.  This is an abscess waiting to happen. However, it is important not to place excessive tension on the suture line or the incision may not heal.  Tacking down tissue layers can be used.  If this is not possible, use of a drain for 3 to 5 days following the procedure is recommended.

Post-procedural Care

It is required that animals be cared for after procedures to ensure their full recovery. Post-procedural care for dogs, cats, swine, sheep and goats is provided by RAR. Post-procedural care for other animals such as small animals, nonhuman primates and in some circumstances, sheep, may be provided by the investigator under RAR supervision, or by RAR if arranged.

Projects outside of the core Minneapolis facilities must make arrangements to either have RAR provide post-op care service if practical or do the care themselves under guidance and oversight from RAR. In either case, the principal investigator should budget accordingly when planning the project. Post-operative care is not included in routine husbandry procedures or the per diem fees.

Post-procedural care includes the following:

1. Monitoring anesthetic recovery

• Someone must be present with any animal recovering from anesthesia until that animal is able to hold itself in a sternal position (on its chest, able to hold its head up). This includes rodents and rabbits
• Endotracheal tubes should be kept in place as long as possible; they must be removed when the animal begins to chew or swallow
• Ability of animal to maintain normal physiology such as body temperature and fluid balance should be assessed.

• 2. Monitoring post-procedural complications 
• Provide analgesia for any procedures with potential for pain or distress
• See standard RAR post-operative analgesia regimens for pigs, sheep, dogs
• Administer antibiotics to prevent post-procedural infections
• Monitor incisions for swelling, exudate, pain or dehiscence
• Monitor catheters & devices
• Monitor for procedure-related complications such as organ failure, thrombosis, ischemia


3. Maintaining records of care given.  These records must include a daily assessment and treatments given.  Other items that could be included in the record are anesthetic agents and time administered, intra-operative assessments and recovery observations.  Post-operative records are required by the USDA on all animals except rats and mice, and must be readily available for review.  Records on rats and mice may be somewhat abbreviated, and can be included as part of research data collected, but should also be available for review.  For more information, contact RAR at 4-9100.

Antibiotic Guidelines

General Recommendations

• If antibiotics are being used, they should be administered before surgery so that they are in tissues when the surgeon is. 
• An appropriate antibiotic should be administered at an adequate dose at the recommended frequency to minimize the development of resistance. 
• Antibiotics should not be used in place of surgical asepsis and good tissue handling techniques. Tissue trauma contributes to post-operative infections.

Selection

If a culture and sensitivity is not available, select antibiotics based on probable organism and probable sensitivity. For example, normal skin flora are usually Gram +, so for a a skin incision, select something with a Gram + spectrum, e.g., amoxicillin/clavulanate or a potentiated sulfonamide. If GI surgery is performed, an antibiotic with a Gram - spectrum is more appropriate, e.g. an aminoglycoside or ceftiofur. Indwelling catheters tend to become infected with skin or fecal contaminants, including anaerobes. Thus a broad spectrum and anaerobic spectrum is required, e.g. amoxicillin/clavulanate, ceftiofur or ticarcillin. Pseudomonas is an opportunist with a high likelihood of a multiple antibiotic resistance phenotype. An extended spectrum penicillin (ticarcillin), a fluoroquinolone (enrofloxacin) or an aminoglycoside (amikacin) may be necessary.

Combinations

Antibiotic activity is classified as being either bacteriostatic (inhibits cells from dividing) or bacteriocidal (kills bacteria even if they are not dividing).  In general, combining two bacteriostatic drugs results in additive effect, combining two cidal drugs results in synergistic effect. Combining cidal and static agents can result in impairment of bacteriocidal activity. If you are treating a specific infection, select two drugs with activity against the organism in question. If you are looking for broad spectrum activity, select drugs with complementary activity, eg. penicillin and an aminoglycoside, or enrofloxacin and clindamycin.

Dosages

Dosages for antibiotics and a description of their activities are listed in RAR's drug formulary.