STANDARD PROTOCOL

Protocol for Obtaining a Surgical Liver Biopsy from Birds

 

USGS, BIOLOGICAL RESOURCES DIVISION                             MODIFICATION DATE: January 10, 2001

ALASKA SCIENCE CENTER - BIOLOGICAL SCIENCE OFFICE                                                                                 

1011 EAST TUDOR ROAD                                 IACUC APPROVAL DATE:________________________

ANCHORAGE, ALASKA 99503                                                              

PROCEDURE TITLE: Liver Biopsy of Birds

SCOPE: This protocol applies to the surgical procedures required to obtain liver biopsies from waterfowl and sea birds.

PRINCIPLE:  Procedures established for obtaining liver biopsies from pet birds usually describe endoscopic biopsies (Altman et al. 1997; Lierz 2000; Nordberg et al. 2000; Rosskopf and Woerpal 1996).  These methods require expensive equipment, are cumbersome and are not suited for field use on wild birds.  The following procedure has been tested for use in the field, and is intended to produce a sample large enough (up to 0.5 g) to allow analysis of cytochrome oxidases.  When such a large sample is not needed, consideration should be given for obtaining a sample by the use of a Tru-Cut̉ biopsy instrument or by a fine needle aspirate.  In galahs (Eolophus roseicapillus), 6% or 18% hepatectomy did not elevate serum bile acid levels or liver enzymes (above those seen with celiotomy alone), and galactose clearance tests were delayed by 18% but not 6% hepatectomy (Jaensch, et al. 2000).  Liver weight as a percentage of body weight was restored to its original value at 7 days after 6% (but not 18%) hepatectomy.

PROCEDURES:

  1. Two people are necessary to obtain the sample: a surgeon and an anesthetist.  Because of the lability of cytochrome oxidases, a third person is required for the immediate handling of the sample.
  2. Standard aseptic surgical technique will be practiced. The surgeon will wear sterile gloves, a hat, and a surgical mask.  The surgical site will be prepared as for any surgical procedure, including plucking feathers, skin disinfection (using povidone iodine or chlorhexidine) and the use of a sterile drape.
  3. Surgical instruments should be sterilized in an autoclave with a minimum of two layers of packaging and stored in a dry place. 
  4. Position the bird on the surgical table in dorsal recumbancy with the legs extended and the wings folded. An insulated (bubble-wrap or foam pad) cover for the surgical table should be used to retard heat loss.  Because regurgitation of crop contents is a frequent problem, birds should be intubated.  Birds should be placed on an elevated platform with a sloped ramp, positioning the bird’s head on the ramp so that it is lower than the body, to avoid aspiration of crop contents.
  5. A single intramuscular dose of ketoprofen at 2 mg/kg is given during or immediately following induction for presumptive post-operative analgesia.
  6. Isoflurane gas anesthetic is administered to the bird by facemask on a non-rebreathing circuit. Induction is at 4-5% isoflurane; maintenance is at 1-2% isoflurane in oxygen or at a level found necessary for a given species and a given individual. Maintenance concentrations of isoflurane may vary depending on the individual bird and environmental variables.  The bird is intubated with a cuffless tube or with a cuffed tube without inflating the cuff.  A protective ointment may be used in the eyes to prevent drying of the cornea.  Once the abdominal air sacs of a bird are opened, respiration can occur partially through the surgical incision, which may require a higher setting on the vaporizer to compensate.  Once the incision is closed, the vaporizer setting may need to be reduced. If oxygen is not available, compressed air (optimum: >50 psi; minimum: 30 psi) can be used to drive the vaporizer.
  7. An alternative anesthetic protocol is the intravenous administration of propofol in combination with a local anesthetic block at the incision site (Machin and Caulkett 1998, 2000).  Propofol does not contain a preservative and supports bacterial growth, so every effort must be made to maintain the stock vial in a sterile state.  Only new needles may be placed into a vial of propofol.  Opened vials of propofol should be kept refrigerated if possible.  Open vials should be discarded if aseptic technique is broken, or within 24 hours of being opened.  A 25 gauge 3/8 in. butterfly catheter is placed into the tibiotarsal vein (alternatively, a 21 gauge, 1 in. butterfly catheter may be placed into the jugular vein.  The catheter is taped in place.  Induction of anesthesia is accomplished by delivering a slow bolus (over 1 min) of 10 mg/kg propofol.  Additional boluses of 1-2 mg may be given to attain induction and to maintain a surgical plane of anesthesia.  All birds must be intubated, and ventilated with a bird AMBU bag.  The incision site and the antenna exit site are infiltrated with 2 mg/kg of a 0.5% solution of bupivicaine, or of lidocaine.  Mortality of male eiders (spectacled and king) has occurred with propofol used as an anesthetic.  Until further information is obtained, propofol should be used only with the greatest caution in male eiders.
  8. Anesthesia is monitored by use of a respiratory or cardiac monitor, or both.  Doppler ultrasound devices or esophageal stethoscopes are the preferred monitors.  An ECG is highly recommended.  Manual palpation of a tibial or brachial arterial pulse can also be used, but is less preferred to an attached continuous electronic monitor.  Respiration is monitored by observation of movements of the bird and the ventilation bag.
  9. Body temperature is monitored with an electronic thermometer with the sensor placed either well into the esophagus or in the cloaca.  The desired temperature range during anesthesia and surgery is 100º to 105º F. The bird should be warmed or cooled to maintain this range.  Additional heat can be supplied to a cold bird by placing bags of warm water on the ventral surfaces of the wings or, ideally, by the use of a radiant heat source located above the bird.  Body temperature can be reduced by removal of external heat sources and by wiping the feet with alcohol or cold water.
  10. Respiration is monitored and mechanically supported when spontaneous breathing is less than one breath per minute.  A minimum of two ventilations per min is made with a bird AMBU bag or with a ventilation bag.  Avoid excess assisted ventilation to avoid delaying a return to spontaneous breathing.
  11. The surgical site is between the distal end of the keel and the conjuncture of the distal ends of the pubic bones, palpated through the abdominal wall.  The feathers are plucked from the site.  An area 1 cm on either side of the incision site should be plucked free of feathers.  The feathers around the site are taped back with pieces of microporous tape.  The site is swabbed twice with povidone-iodine or benzalkonium chloride solution.  A sterile fenestrated drape is placed over the surgical site.
  12. The skin is incised along the ventral midline with a No. 11 or No. 15 sterile blade. The subcutaneous layer and fat are sharp dissected. Once the muscular abdominal wall is reached, the linea alba is identified. The linea alba is seized with a forceps and lifted to permit penetration of the abdominal wall with a blade. The linea alba is then sharp dissected with blade or scissors, avoiding the viscera, to a length of about 2 cm.
  13. The caudal edge of the liver is located in the abdomen.  Occasionally the liver is located too far rostrally to permit easy access.  In those cases, the surgeon should pick up the bird, keeping his hands on the sterile side of the drape, elevate the bird’s head, and shake the bird gently.  The liver should drop down into view. 
  14. A piece of the caudal edge of the liver is isolated using a curved mosquito forceps.  The liver should fill the entire arc of the forceps.  The jaws of the forceps are closed, but not locked, crushing the tissue.  The isolated piece of liver is then cut free by passing a scalpel blade along the inside curve of the forceps.  The cut piece of liver is then placed on a sterile sponge and dropped onto the surgical table outside the sterile field, for the sample processor to pick up.
  15. The surgeon maintains the crush of the cut edge of the liver for about 10 seconds.  Then the forceps is gently released, observing for hemorrhage.  If bleeding is observed, the forceps are gently applied to the bleeding part and pressure is held for another 10 sec.  The forceps are then gently removed and the cut edge observed for bleeding.  Gel foam can be applied to the cut edge of the liver if continued bleeding is a problem.
  16. The surgical incision is closed in two layers using 3-0 braided absorbable sutures on a cutting needle. The linea alba is closed using a simple continuous pattern and the skin is closed using either a simple continuous or simple interrupted pattern.
  17. The drape is removed and the vaporizer is turned to zero. Oxygen supplementation should continue until the bird recovers. Additional procedures such as obtaining a blood sample or banding may be done during this period. The bird should be kept warm by holding it wrapped in a towel until it is fully recovered. If dehydration is a problem, subcutaneous fluids can be administered.
  18. Following recovery, the bird should be placed in a cage or kennel for at least one hour prior to release.  Birds should be released only when they are alert, able to maintain head and body position, and react to human handling.  Birds that do not respond should be carefully inspected and supportive care (heat source, gastric intubation of water and electrolytes, etc.).

REFERENCES:

 

Altman,R.B., S. L. Clubb, G. M. Dorrestein, K. Quesenberry. 1997. Avian Medicine and Surgery. W.B. Saunders Co. Philadelphia.

Jaensch,S.M.; L. Cullen, and S. R. Raidal. 2000. Assessment of liver function in galahs (Eolophus roseicapillus) after partial hepatectomy: a comparison of plasma enzyme concentrations, serum bile acid levels, and galactose clearance tests. J. Avian Med. Surg. 14(3):164-171.

Lierz, M. 2000. Endoscope-guided biopsy in birds. Exotic DVM 2.3:17-25.

Machin, K. L., N. A. Caulkett. 1998. Investigation of injectable anesthetic agents in mallard ducks (Anas platyrhynchos): a descriptive study.  J. Avian Med. Surg. 12 (4): 255-262.

Machin, K. L., N. A. Caulkett. 2000. Evaluation of isoflurane and propofol anesthesia for intraabdominal transmitter placement in nesting female canvasback ducks. J. Wildl. Dis. 36(2):324-334.

Nordberg,C. R.T. O’Brien, J. Paul-Murphy, B. Hawley. 2000. Ultrasound examination and guided fine-needle aspiration of the liver in Amazon parrots (Amazona species). J. Avian Med. Surg. 14(3):180-184.

Rosskopf, W.J.Jr. and R. W. Woerpel. 1996. Diseases of Cage and Aviary Birds. Williams & Wilkins. Baltimore.