IMAGING INSIDE TORTOISES

Sharon Redrobe BSc (Hons) BVetMed CertLAS DZooMed MRCVS

RCVS Recognised Specialist in Zoological Medicine
Clinical Associate Professor of Zoo, Wild and Exotic Animal Medicine, School of Veterinary Medicine and Science, University of Nottingham
Director of Life Sciences, Twycross Zoo, Twycross Zoo East Midland Zoological Society
Email: sharon.redrobe{at}nottingham.ac.uk
www.twycrosszoo.org
Presented to the BCG Symposium at the Open University, Milton Keynes, on 13th March 2011.

Ultrasonography and radiography

These are complementary imaging techniques. In rare cases one technique is used instead of another but more commonly both modalities are used to maximise diagnostic information. Positioning is similar to the dog and cat for most procedures.

Ultrasonography is an excellent modality for differentiating between fluid and soft tissues and is therefore the diagnostic test of choice to investigate lesions of the bladder, heart and uterus. Radiography requires contrast material to fully explore these organs. Radiography is useful in determining the size, shape and position of organs within the body and relative to each other as large areas of the body can be imaged. Ultrasonography only allows imaging of a small area of the body at a time but allows visualisation of the internal organ structure. Ultrasonography is a dynamic modality and is therefore useful for assessment of cardiac movement and blood flow using the Doppler facility. Ultrasound waves are blocked by gas and therefore the ultrasonography of the abdomen of small herbivores is greatly hampered by the pockets of gas present. Artefacts and over-diagnosis are common in ultrasonography. It is vital therefore that the ultrasonographer is familiar both with the normal anatomy of the species under examination as well as with the basic practice of ultrasonography. Radiography is a much more familiar imaging technique to most clinicians and the images once taken can be read by others. Interpretation of the ultrasound image requires knowledge of where the probe was placed on the animal, moving images being more easily interpreted than still images, making second opinions of ultrasonographic findings difficult.

Radiography

Correct positioning is important. Animals can be taped down, or radiographed through a box or bag if not sedated. Three views are typically required:

• Dorsoventral view – take care; a healthy animal can move very quickly off the table! Take exposure between expiration and inspiration. Placing animal on a raised column with the feet off the table can aid restraint.
• Lateral view – tilting chelonians onto their side distorts the diaphragm and lungs, thus horizontal beam required; centre beam on 6-7th marginal shield at right angles to vertebral column.
• Cranio-caudal – useful for contrasting two lung fields. Centre horizontal beam on nuchal shield, head, neck, limbs. General anaesthesia required for optimal positioning of extremities.

Ultrasonography

The diagnostic potential of radiology in chelonia is limited by the bony plastron and carapace. The skeleton and pulmonary fields are readily imaged, as are calcified eggs and uroliths (bladder stones). Soft tissue structures are not apparent unless gas-filled or abnormally calcified. The investigation of gastro-intestinal tract abnormalities by radiography requires the use of contrast media (Taylor et al., 1996). Coelioscopy (a fine camera is inserted into the body), magnetic resonance imaging (MRI) or computed tomography (CT which is 3D x rays) are other alternatives. Coelioscopy is invasive and requires full general anaesthesia, MRI usually requires some as it takes a long time, but CT is quick and requires no sedation. Coelioscopy does allow for biopsies to be taken of internal organs for analysis.

The advent of ultrasonography has made possible the non-invasive, relatively safe assessment of soft tissues in many species of reptile. In many cases chemical restraint is not necessary. Ultrasonography is the major imaging tool for obstetrical work in reptiles, being useful in infertility assessment, determination of the number of follicles and relating the timing of ovulation to egg laying (Stoskopf, 1989). Some aspects of chelonia ultrasonography have been reported by Penninck and others (1990).

The chelonia species are usually examined without chemical restraint. The animal is held in a dorsal horizontal position with the feet held away from the table to prevent movement. For all but the largest species, one person restrains the animal whilst another performs the ultrasound scan. Aggressive turtles/terrapins may require anaesthesia to prevent damage to the transducer or operator. A short acting agent e.g. propofol 15mg/kg intravenously is useful in these cases. This has proved rarely necessary.

The shell and underlying bony plates of chelonia restrict the positioning of the transducer to the ‘acoustic windows’ in the inguinal, pre-femoral, axillaryand cervical regions where the transducer may be placed in contact with soft tissue. Cervical and pre-femoral windows are used in most circumstances. A 5MHz sector transducer may be used with larger species. The 7.5 and 10MHz sized sector probes are useful in smaller patients, i.e. those weighing less than 500g. The resolution is greater but penetration of the beam less with the greater frequency probes. Acoustic coupling gel is applied liberally to the skin and transducer. If the physical size of the transducer head prevents placement between the bony bridge, then an acoustic stand-off is required. This can be achieved by using a water-filled surgical glove placed between the transducer and the patient’s skin. Alternatively, the animal and scanner are immersed in a water bath. The transducer is held angled towards the soft tissue acoustic window. Transmission of the ultrasound beam is satisfactory up to a distance of 2-3cm through the water. Adult red-eared terrapins and Mediterranean tortoises are imaged using a 5 or 7.5MHz transducer without a stand-off. When an image is obtained, the focus and depth are adjusted to enhance the image quality. Species scanned include Hermann’s tortoise (Testudo hermanni), Greek or spur-thighed tortoise (T. graeca), American box turtle (Terrapene spp) and red-eared terrapin or slider (Pseudemys scripta elegans).

A standard procedure for ultrasonography ensures that each system is imaged. We begin by imaging the heart from the cervical window. The coelomic organs are then imaged from the pre-femoral window. The usual order is bladder, liver, gonad. To access the cervical window, the head and foreleg must be fully extended and pulled to the side. This is the preferred site to image the heart. The heart is imaged on the midline in the dorsal third just caudal to the scapulae. This distance may be estimated from a dorso-ventral whole body radiograph. The rear limb fossae are the site for ultrasonographic examination of the caudal coelom (body cavity). This is the site preferred to image the liver, gastrointestinal tract and genito-urinary tract.

• The bladder is imaged from the pre-femoral window and by angling the transducer towards the midline. The urine filled bladder is seen as an anechoic oval structure. Urates may be detected as hyperechoic specks (brighter echoes) floating within it. Uroliths are imaged as hyperechoic structures within the bladder. The bladder wall is usually only detected in cases of bladder wall thickening caused by such chronic irritation.
• The liver is readily imaged through the pre-femoral fossae, and is cranial to the bladder, spanning the midline. Depending on the size of the animal and transducer, the heart may be imaged using the liver as an acoustic window. This does not give such a good image as using the cervical window, however. The liver has a uniformly homogenous echotexture, similar to mammals. Portal vessel and hepatic vessels can be seen, the walls of the latter appearing less echogenic. The gall bladder can be seen on the right side as an anechoic structure. Liver abscesses and scarring are detected as a change in the homogeneity of the image. Hepatic lipidosis is seen as a diffuse increase in echogenicity. Liver abscesses are commonly reported at post-mortem in aged chelonia. These lesions are readily detected using ultrasound when they appear as discrete disruptions to the architecture of the liver. Distinction between abscess and neoplasia does, however, require histopathology. The finding and extent of hepatic lipidosis in the anorexic chelonian may serve as a prognostic indicator.
• The testes are located cranial and ventral to the kidneys. They appear uniformly echogenic and slightly more hypoechoic when compared with the adjacent kidneys in chelonia.
• The ovaries in chelonia are paired elongated organs attached to the peritoneum on either side of the dorsal midline anterior to the pelvic girdle. The expanded tissue of the ovaries ensures that enlarged follicles may be imaged by ultrasonography as they are positioned between other organs, e.g. oviducts or intestinal loops. Care must be taken to differentiate follicles from loops of intestine; as the transducer is rotated through 90 degrees, the follicles will retain a spherical appearance whereas the loops of bowel will appear as tubes. The follicles are more easily imaged against the background of a large fluid filled bladder. When ovulated, the hyperechoic yolk is seen centrally surrounded by the dark hypoechoic albumin. As the shell is deposited, the dense white hyperechoic margin of yolk is seen surrounded by the dense fibrous layers of egg shell increasing the echogenicity of the egg (Kuchling, 1989).

The accurate estimation of the number of follicles or eggs is not possible in those species which mature large numbers of follicles. Ultrasound can penetrate thin-walled eggs of some species, e.g. red-eared terrapins, to detect the inspissated (congealed) contents within retained eggs. This has proved a useful guide in determining the age of the egg and hence in diagnosing dystocia. Eggs which are in the distal tract may be imaged through the inguinal fossa (between the tail and legs).

Ultrasound has been used to detect, count and measure the ovarian follicles and the oviductal eggs in sea turtles (Kuchling, 1989). In most species the character of the ovaries can be assessed, eggs may be detected in the oviduct, and maturing and ovulating follicles may be monitored to allow timing of matings to be estimated. The pregnancy may be monitored and related to nesting behaviour and the correct timing of oviposition.

Recent work in Kemps’ Ridley sea turtle (Lepidochelys kempi) found that the resolution using a linear scanner was better than a sector scanner at distinguishing follicles (Rostal et al., 1990). However, the size of many ‘pet’ species precludes the use of a larger linear array transducer. The resolution using our system as described has proved adequate.

Case example

A six-year-old female red-eared terrapin (P. s. elegans) weighing 680g was housed with an adult male. The tank consisted of a body of water and a solid basking platform. The male had been observed courting the female. The female was scanned using a 10MHz transducer every three days during this period of courtship. The follicular activity was followed. Follicles appeared as multiple hyperechoic spheres on the ovary. The oviductal egg was seen surrounded by dark hypoechoic albumin. The number of eggs produced was confirmed by radiography. The female was then provided with a suitable nest site and she laid the full clutch of eggs three weeks after follicular activity was first recorded.

Echocardiology has proved rewarding in chelonia. The heart can easily be imaged in animals weighing over 300g. The ventricular and atrial walls, atrioventricular valves and main coronary vessels can be imaged. Abscesses can be detected in the major vessels and associated on the valve leaflets. The relative thickness of the myocardium allows investigation by M mode ultrasonography in medium sized specimens, e.g. Greek (T. graeca) and Hermann’s (T. hermanni) tortoises weighing over 500g. The blood flow pattern and velocities can be measured using the colour Doppler display and spectral analysis of flow direction and velocity. At present, gross lesions such as major valvular incompetence can be noted.

The presenting signs associated with cardiac disease include peripheral oedema, ascites (fluid in the body cavity), weakness and dyspnoea. Echocardiography may prove rewarding in these cases and should be performed to investigate these symptoms. At the Royal (Dick) School of Veterinary Studies a number of animals have had the Doppler colour flow technique performed and M mode values assessed. When sufficient baseline values have been established, these parameters may prove useful in the detection of valvular insufficiencies, stenoses etc.

Ultrasonography is useful in the location of a lesion to a specific organ, investigating disease of the liver and other soft tissue organs, the detection of ovarian activity and pregnancy, plus the detection of some cardiac abnormalities. More work in elucidating the ‘normal’ will aid in the use of this modality for the ‘abnormal’ patient. These preliminary studies indicate the great potential of ultrasonography in the management of the chelonian patient.

Endoscopy

Appropriate general anaesthesia is required for invasive endoscopy. Even oral/respiratory tract endoscopy requires general anaesthesia in most if not all reptiles to permit adequate restraint and prevent damage to the animal and/or endoscope.

Coelomic organ endoscopy

1. With the chelonian in lateral recumbency, an incision is made in the prefemoral fossa. Insufflation is required to space the organs and identify them by visualisation.
2. The tips of graspers or manipulating forceps can be used to push aside organs (particularly the gastrointestinal tract) to navigate around the coelom.
3. It is important to be familiar with the location of the coelomic organs so that the incision can be made in an appropriate place.
4. It is possible to visualise and identify the heart, lungs, liver, adrenal, gonads (testes or ovaries), oviduct in females, kidney, ureters (with urates passing down), gastrointestinal tract, pancreas and spleen.
5. It is possible to visualise and identify the heart, lungs, liver, adrenal, gonads (testes or ovaries), oviduct in females, kidney, ureters (with urates passing down), gastrointestinal tract, pancreas and spleen.

Chelonian lung endoscopy

1. Direct access to a lung lesion identified on radiography can be gained by drilling through the shell at the appropriate site.
2. The endoscope is introduced and a biopsy and/or cytology sample/flush is conducted.
3. The bony defect is sealed with bone wax or dental acrylic and will heal in a few weeks.
4. Aquatic chelonia should be kept out of water for 2-7 days.
5. In the case of obligate aquatic turtles, e.g. soft shelled turtles which cannot be kept out of water, the defect should be additionally closed with surgical glue so that the animal can be immersed as soon as it has recovered from general anaesthesia and then observed for signs of air escaping from the defect on emersion before being released to the owner.

Cloaca

Indications – tenesmus (straining), oviduct pathology, haematuria, prolapse.

1. The reptile is placed in ventral recumbency, with the tail elevated and pushed cranially.
2. A giving set tube (attached to a bag of warm saline) is introduced into the cloaca with a scope, or saline is attached to the obturator port of the scope.
3. The cloaca is closed around the scope, so that fluid instilled remains inside and facilitates full visualisation.
4. The cloaca is entered and the coprodeum, or deepest part of the cloaca, will be found as an inner sphincter, often with faeces present.
5. The entrance of the ureters and oviduct may also be visualised.

Biopsy Collection

Via the coelomic entry, the surgeon should be able to gather biopsies from kidney, lung and liver.

References

Kuchling, G. (1989). Assessment of ovarian follicles and oviductal eggs by ultrasound scanning in live fresh water turtles (Chelodina oblonga). Herpetologica 45(1): 89-94.

Penninck, D.G., Stewart, J.S., Paul-Murphy, J. & Pion, P. (1991). Ultrasonography of the Californian desert tortoise (Xerobates agassizi): anatomy and application. Vet. radiography 32(3): 112-116.

Rostal, D.C., Robeck, T.R., Owens, D.W. & Kraemer, D.C. (1990). Ultrasound imaging of ovaries and eggs in Kemp’s ridley sea turtles (Lepidochelys kempi). J. Zoo and Wildlife Medicine 21(1): 27-35.

Stoskopf, M.K. (1989). Clinical imaging in zoological medicine: A review. J. Zoo and Wildlife Medicine 20(4): 396-412.

Taylor, S.K., Citino, S.B., Zdziarski, J.M. & Mitchell Bush, R. (1996). Radiographic anatomy and barium sulfate transit time of the gastrointestinal tract of the leopard tortoise (Testudo pardalis). J. Zoo and Wildlife Medicine 27(2): 180-186.

Editor’s note: the author is the winner of the 2009 British Chelonia Group’s Oliphant Jackson Award for outstanding contribution to tortoise medicine for work on ultrasonography in tortoises, including the paper Redrobe, S.P. & Scudamore, C. L. (2000). Ultrasonographic diagnosis of pericardial effusion and atrial dilation in a spur thighed tortoise (Testudo graeca). Vet. Rec. 146(7): 183-185. Summary published in Testudo 7(1), 2009.

Testudo Volume Seven Number Three 2011.

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