(See also Necropsy of Mammals (Techniques Overview) )

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• Post mortem examination (i.e. necropsy) is an extremely valuable tool in disease investigation and management. It is important to approach each carcass with an open mind, not assuming that the cause of death is known, even if there are obvious external lesions or a known on-going disease problem.
• N.B. Examination of embryos and dead-in-shell chicks may be useful as well as examination of adult animals.
• In performing a necropsy or post mortem examination, it is important to:

• have a systematic approach, whether head-to-tail, system by system (digestive system, respiratory system etc.) or any other.
• recognize the normal anatomy, normal appearance of organs/tissues and anatomical variation between species.
• accurately describe lesions/abnormalities.
• keep accurate records, including a unique identifying number for each carcass and samples from that carcass.
• avoid the use of non-standard abbreviations in permanent records.
• preserve samples (tissues, parasites etc.) for future reference/research

• Suitable dissection instruments, sterile culture swabs, clean slides for impression smears, appropriate sterile containers, fixation medium (buffered formalin, also 70% alcohol) should be available.
• N.B. In the event of a die-off it is important to examine a number of individuals, representative for the range of species affected, and to remember that more than one disease process may be acting at any one time and that the major cause(s) of death may change during a prolonged die-off.
• The results of the post mortem examination should be used in conjunction with the history of the bird or birds and assessment of the environment.  (See History and Documentation - General and Environmental Assessment - General)

(B11.11.w20, B15, B36.2.w2, B36.3.w3)

• Knowledge of normal anatomy is useful e.g. long coiled trachea in some swan species and long tracheal loop sub-cutaneous over sternum of  Anseranas semipalmata - Magpie goose.

• Chapter 2 - B36 Field Manual of Wildlife Diseases - Specimen Collection and Preservation
• Chapter 3 - B36 Field Manual of Wildlife Diseases - Specimen shipment

GENERAL:

• Record the age and sex (if known), any identifying number, bands, tags etc. Retain any rings, microchips etc.
• Examine the inside of the container/wrappings the carcass has been presented in, noting contaminants (e.g. mud, oil) and possible external parasites such as leeches, lice or mites which may have left the host.
• Weigh the carcass.
• Examine the external surface of the carcass. Note whether the carcass is fresh or decomposed and whether it has been frozen, also whether it is intact or scavenged, and if scavenged to what degree. Permanent written records should indicate the following:

• General appearance of carcass and plumage  - e.g. wet, muddy, oiled, clean, bloody, covered in salt; feathers intact, damaged or missing. If contaminated, note extent and areas affected.
• Presence of parasites - leeches (particularly around eyes and nares), lice, ticks etc.
• Body condition - prominence of keel
• Examine: bill (outside and inside), mouth, external nares, eyes, ears, head, neck, body, uropygial gland, wings, legs. Check for discharges from body orifices, scouring, abrasions, lacerations, fractures, dislocation.

• Radiography may be indicated to detect e.g. fractures, radiodense foreign bodies (particularly lead shot in the gizzard, evidence of shooting, fishhooks), metabolic bone disease.
• Preparing for internal examination: Wet feathers thoroughly with soapy water or disinfectant in water to minimize feather debris.
• N.B. tissue samples, swabs for culture and impression smears should be taken as tissues are examined and before they become contaminated.  Tissue to be fixed should be no more than 3-5mm thick, and thinner if congested, although whole lung may be fixed.

EXAMINATION BY SYSTEMATIC DISSECTION:

• Examination of head: cut through sinuses, cut through corner of bill, examine hard palate, tongue. Cut skin over head and peel back, cut through cranium on both sides from foramen magnum forwards, keeping scissors or shears perpendicular to surface to minimize damage to brain, and lift off. Examine brain in situ, note any haemorrhage (N.B.  agonal bleeding may produce blood in cranial bone, but haemorrhage in meninges or brain tissue should be considered significant). Cut brain sagittally, retain samples for toxicology. Eyes may be removed with care and fixed in a preservative preferred by the histopathologist, e.g. Bouin's or Zenkers solution.
• Slit the skin from the vent to the bill in the ventral midline: nick the skin over the breast muscles in the midline, continue the incision rostrally (forwards) to the bill and caudally (backwards) to the cloaca (vent), taking care not to damage underlying tissues.
• Reflect the skin from the neck, chest and abdomen , noting amount of subcutaneous fat, amount of breast muscle present, thymus in young birds, colour of subcutaneous tissues, bruising, haemorrhage or other lesions, condition of external surface of trachea and oesophagus, distension/impaction of oesophagus, presence of haemorrhage, pale areas or e.g. 'rice grain' lesions in breast muscles, presence of haemorrhage in neck muscles.
• Incise over major limb joints, examine state of synovial fluid, joint surfaces and tendon sheaths. Incise leg muscles, note colour (e.g. pale areas), any haemorrhage. Leg bones may be checked for whether they may be bent or easily broken.
• Remove sternum and rib cage: Incise transversely (across the body) just below the sternum, lift the sternum up wards and, noting the condition of the air sacs, cut through the rib cage and coracoid on either side using shears.
• Note any gross abnormalities (e.g. gross haemorrhage), and examine pericardial sac, thyroid and parathyroid glands (at thoracic inlet, near common carotid arteries). Dissect out the heart, leaving a portion of the major blood vessels intact.  Examine the heart: overall shape,  pericardium, fluid in pericardial sac, epicardial surface - any haemorrhages, presence of worms visible under surface, thickness of ventricle walls, presence and amount of blood in ventricles, state of valves, colour of muscle: haemorrhages or pale areas within muscle or on endocardial surface, cut along major vessels, check walls of vessels for  presence of plaques, calcification. [Normal:- pericardial sac clear/translucent, containing trace of clear fluid. Epicardial surface clean, glistening, heart contracted, triangular. left ventricle wall two to three times as thick as right ventricle wall, little or no blood in left ventricle, small amount of blood in right ventricle. Valves smooth and shiny]. Cultures and smears may be prepared from heart blood.
  • A scintillating sheen of urate crystals may be visible over the viscera in a bird with visceral gout. (B14)
• Examine liver in situ, note size, shape, colour, any haemorrhage on surface or free in body cavity, whether surface is intact or split, presence of pale areas, whether lesions are flush with surface or protruding or shrunken, whether edges of lobes are sharp (normal) or rounded (enlarged).  Remove liver, carefully cutting connections with other tissues. Cut through liver several times, note colour variations, size and shape of any lesions (pale areas, haemorrhages, abscesses). Note size of gall bladder. Take tissue/swab for culture, impression smears from cut surface, tissue for histology and toxicology (e.g. lead levels in waterfowl) as appropriate.
• Examine lungs in situ, note colour. Cut through lungs, noting any presence of water, froth, blood, fungal infection, abscesses.  Using sharp scissors, cut along whole length of trachea (cut along both sides required, as tracheal rings are springy) from mouth, and major bronchi, noting presence of fluid, blood, fungal plaques, necrotic lesions, parasites (tracheal flukes, gapeworm). N.B. if drowned, may be water in air sacs and lungs, or just lungs if there has been a delay in examination.
• Examine gastro-intestinal tract in situ, note any distension, discolouration, haemorrhage or lesions on serosal surface, size, shape and colour of spleen (normal small and flat in waterfowl). Note presence and type of any fluid within body cavity. Tie off oesophagus at top of neck, cut above this, remove whole of gastro-intestinal tract.
• Examine kidneys, adrenals, and reproductive tract in situ, including size and colour of kidneys, presence of haemorrhage or pale areas (e.g. with renal coccidiosis, presence of chalky material if uric acid build up. Examine gonads: size, whether ovary is active (follicles present and growing) or inactive, appearance of oviduct, presence of egg within the oviduct or the abdomen, presence of associated reaction if egg or yolk is present free within body cavity, any haemorrhage. Remove kidneys, adrenals, gonads, incise and examine. 
• Examine whole of gastro-intestinal tract, including pancreas. Open each region (oesophagus, proventriculus, gizzard, small intestine, large intestine, caeca, cloaca). Check for: haemorrhage (N.B. agonal bleeding and post mortem leakage must be distinguished from haemorrhage into gut), ulceration or other lesions, presence of parasites and whether lesions are associated with these, presence and appearance of ingesta, including recognizable food items in the oesophagus and gizzard, and any foreign bodies. Remove any parasites, preserve for identification.  Fresh samples of contents and mucosal scrapings of duodenum, ileum, caeca may be examined microscopically for parasitic ova and coccidial oocysts, also smears may be Gram stained. Gizzard: Flush out gizzard contents into white bowl,  examine for presence of lead shot and other foreign bodies, note colour of gizzard lining and whether it is normally adherent to the underlying tissue and whether it can be peeled out.

(J34.14.w1, B11.11.w20, B14, B15, B36.2.w2, P18.w1)

HISTORY: Egg history is an important part of egg necropsy and the following details should be recorded:-

• Species, identity and previous breeding history of the adult birds.
• Date that the egg was laid.
• Details of incubation:

• Parent, broody or artificial incubation.
• Any known disturbance of incubating birds.
• Incubation records - incubator type, records detailing intended incubator settings and actual temperature, humidity, turning, ventilation etc.
• Whether eggs were stored before incubation and if so details of storage length of time, temperature, turning.
• Period of incubation (normal incubation time for this species, actual time egg incubated prior to necropsy)
• Results of any egg inspections (candling, flotation) during incubation.
• Initial weight of egg, record of weight loss.
• Whether other eggs from this clutch and/or in the same incubator have failed to hatch and if so what percentage.

EXAMINATION OF THE EGG:

• Examine external surface. Note any: blotchiness/discolouration, oozing, cracks, presence of dirt, any identifying number for records, if there is chipping/pipping of shell and whether the bill is visible if pipping has occurred.
• Weigh and measure length and width.   (N.B. small eggs may be associated with salpingitis). Photograph egg.
• Candle the egg. Note whether the egg appears  probably infertile (clear) or probably fertile. Sketch any findings.  - size and position of air space, size and position of embryo if visible, any other findings.
• Open the egg. Clean the shell with 70% alcohol (methanol) and allow to dry. The shell may be opened by removing an elliptical piece of shell over the long axis of the shell (P1.1989.w1, B116.9.w1), or by removing a piece of shell over the large end, i.e. over the air cell (B11.11.w20). Puncture the shell with a sharp pair of scissors, remove a sufficiently large piece of shell to visualize the contents of the egg, and place this to one side. Note shell thickness, state of membranes if dead-in-shell. [Long-axis approach may give better visualization of the yolk and any blastocyst (to distinguish fertile from infertile eggs), and of the position of the embryo, without affecting the yolk or aircell].
• Examine the contents of the egg in situ: describe the contents, including a sketch and/or photograph as appropriate.
• For eggs thought to be infertile: sample for bacterial, fungal, viral culture. Retain contents if required for toxicological examination (store at -20C), otherwise discard.
• For eggs thought to be fertile:

• Culture contents or swabs from different sites (depending on degree of development). Swab sites may include e.g. yolk, amniotic fluid, vitelline membrane. N.B. analysis of findings must take into account possibility of contamination of long-dead or infertile eggs by environmental bacteria.
• Examine in situ for malpositioning. Note particularly position of bill if dead-in shell.
• Remove embryo and associated tissues from egg.  Weigh and measure. N.B. note weight of embryo both including and without yolk sac. Measurements may include e.g. crown-rump length, culmen length, ulnar and tarsal lengths. An estimate of the age of the embryo should be made, although accurate age determination may be difficult as details of development are available for only a few species.and estimates must take account of the normal incubation period for the species and whether the chicks are altricial or precocial (P1.1989.w1).
• Examine embryo as if it were a hatched chick. N.B. a dissecting microscope may be useful, and sketches and photographs taken as appropriate. Note developmental abnormalities such as prognathia, hydrocephalus. Examine the yolk stalk and umbilicus with care. Small embryos may be fixed whole and sectioned. Larger embryos may be dissected, with samples of tissues taken for bacteriology, mycology, histopathology, virology or other tests and the remaining embryo may be fixed (10% buffered formalin, or alcohol or Bouin's solution) and retained. N.B. yolk sac and contents should be separated from other tissues and stored separately to avoid yolk droplets/granules confusing the histopathological findings. Yolk sac may be used for culture, vitamin analysis, toxicological analysis. Embryo, yolk etc. may be frozen for whole-egg toxicological analysis.
• Examine shell membrane, aircell, allantois, amniotic sac and fluid, yolk sac, contents and vitelline circulation.
• N.B. Histological examination may be used to distinguish whether "suspicious material" on the yolk surface or in a severely addled egg is an early embryo or not.
• Histopathological findings may be compromised due to the degree (often considerable) of autolysis, but may be useful both in determining the cause of death and in developing information on the normal histology of avian embryos.
• Toxicological tests which may be carried out include those for chlorinated hydrocarbons, heavy metals and selenium.
• Shell: dry for seven days and weigh.

• N.B. interpretation of the findings may be difficult. Even distinguishing between an infertile egg and an early embryonic death is difficult as eggs may be severely decomposed by the time of examination. Management factors affecting the parents, genetics, egg storage and incubation conditions (temperature, humidity, turning) as well as the effects of toxins, viruses, bacteria and fungi must be considered. Secondary infection is common and usually results in mixed bacterial cultures. Heavy growth of one or two organisms may be more likely to indicate a primary infection.

(B11.11.w20, B116.9.w1, B116.11.w2, P1.1989.w2)

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• Potential hazards to human health of ANY necropsy or post mortem examination must be considered before undertaking examinations.
• Personnel undertaking or attending necropsies or post mortem examinations must be made aware of the  potential hazards to human health.

FOR BIRDS:

• Potential hazards range from toxins on the surface of the animal (e.g. oil) to zoonotic diseases which may be transmitted through cuts, absorbed through mucous membranes or inhaled in the form of dust or aerosols.
• Important zoonotic diseases to consider in dealing with bird carcasses include Aspergillosis, Avian Tuberculosis, Chlamydiosis / Psittacosis, Erysipelothrix Infection, Salmonellosis and Yersiniosis.
• For all necropsies, protective clothing should be worn, particularly disposable gloves (which should be replaced immediately if damaged). A face mask is advisable.
• If possible, necropsy should be carried out inside a protective cabinet
• Any cuts should be washed immediately with a disinfectant soap.

(B11.11.w20, B96.w2)

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