Received: 6 July 2017 Revised: 16 November 2017 Accepted: 19 November 2017

DOI: 10.1111/vru.12599

O R I G I N A L I N V E S T I G AT I O N

Ultrasonographic anatomy of reproductive female leopard

geckos (Eublepharis macularius)

Ophélie Cojean1 ∗ Claire Vergneau-Grosset1 Isabelle Masseau2

1 Service de médecine zoologique, Faculté de

médecine vétérinaire, Université de Montréal, Abstract

Saint-Hyacinthe, J2S 2M2, Québec, Canada
2 Department of Clinical Sciences, Faculté de
médecine vétérinaire, Université de Montréal,
Saint-Hyacinthe, J2S 2M2, Québec, Canada
Correspondence
Isabelle Masseau, Department of Clinical Sci-
ences, Faculté de médecine vétérinaire, Uni-
versité de Montréal, Saint-Hyacinthe, J2S 2M2,
Québec, Canada.
Email: isabelle.masseau@umontreal.ca
Captive leopard geckos (Eublepharis macularius) often present to the exotic clinic for gastrointesti-
nal impactions, follicular stasis, or dystocia. To our knowledge, normal ultrasonographic anatomy
of these lizards has not been described. The objectives of this prospective, anatomic, analytical
study were to develop ultrasound techniques for this species and to describe the normal sono-
graphic anatomy of the head, coelomic cavity, and tail. Eleven, healthy, female leopard geckos were
included. A linear array 13–18 MHz transducer was used. Geckos were sedated and restrained in
dorsal recumbency for coelomic structure examination and in ventral recumbency for head and
tail examinations. Sagittal and transverse images were acquired and authors recorded qualitative
and quantitative ultrasonographic characteristics of anatomic structures. The ventral surface of
the lungs, liver, gallbladder, caudal vena cava, portal vein, ventral abdominal vein, aorta, ovarian
follicles, fat bodies, tail, and brain were visualized in 10 of 11 individuals. In one individual, molt
precluded ultrasonographic examination. The heart, kidneys, urinary bladder, spleen, and pan-
creas were not visualized. The digestive tract was observed in 10 individuals but was too small
to be measured. Findings from the current study could be used as a reference for future studies of
leopard geckos.

KEYWORDS
anatomy, imaging, lizards, ultrasonography

1 INTRODUCTION Leopard geckos (Eublepharis macularius) are emblematic lizards

Ultrasonography is an imaging modality commonly used in mammals
and is increasingly being used in reptile medicine.1 Anatomical differ-
ences between mammals and reptiles include the lack of diaphragm,
the presence of renal portal system, and of two fat bodies located
in the most caudal part of their coelom. It has been postulated that
these fat bodies constitute a reserve of adipose tissue compensating
for the lack of subcutaneous fat in most of their body except in the
subcutaneous tissue of the tail.2 Indeed, the tail of the leopard gecko
serves as an important site of fat storage and similar to other fat-
tailed species, lipid deposits are distributed all along the length of the
tail.2 The ultrasonographic appearance of coelomic organs has been
reported in bearded dragons (Pogona vitticeps),3 green iguanas (Iguana
iguana),4 bosc monitor (Varanus exanthematicus),5 veiled chameleons
(Chameleo calyptratus),6 loggerhead see turtles (Caretta caretta),7
constrictor (Boa constrictor),8 and other boid snakes9 among others.
∗ Present address: Dr. Ophélie Cojean, Exotic Companion Animal Service, Benjamin Franklin
Veterinary Clinic, 38 rue du Danemark, ZA Porte Oceane II, 56400, Brech, France.
commonly kept and bred in captivity due to their small size and per-
ceived limited husbandry requirements. This insectivorous and noc-
turnal species originates from the South–West of Asia.10 Leopard
geckos have been extensively studied for their ability to autotomy
and regenerate their tail11 or for their temperature-dependent sex
determination.12 Although leopard geckos have been bred for dual
purposes of companion and laboratory animals for decades, there is
currently no study describing the normal ultrasonographic anatomy
of this lizard. Common diseases of leopard geckos, such as gastroin-
testinal impactions, follicular stasis, dystocia,13 and neoplasia could
potentially be diagnosed by coelomic ultrasound as described in other
gecko species.14 In addition, although more rarely described, diseases
of the central nervous system such as xanthomas,15,16 intracerebral
neoplasms,17 or hydrocephalus18 occur in reptiles including leopard
boa geckos. These are possibly under-diagnosed because the diagnosis so
far requires magnetic resonance imaging, but ultrasound of the brain
could be a good alternative. Imaging the central nervous system of
leopard geckos would also allow to investigate and rule out large
cerebral lesions in individuals presented with neurologic signs. For
instance, leopard geckos may present with seizures due to metabolic

Vet Radiol Ultrasound. 2018;1–12. wileyonlinelibrary.com/journal/vru 

c 2018 American College of Veterinary Radiology 1
2 COJEAN ET AL .
disorders such as hypocalcemia19,20 or due to congenital neurologic
disorders,21 such as geckos of the morph called Enigma frequently
affected by seizures. Ultrasound examination of the brain would be a
noninvasive technique to rule out large intracerebral lesions in these
cases. Ultrasound examination of the tail could be useful to evalu-
ate hemipene lesions, as reported in other lizards,22 as well infectious
skeletal lesions which often affect vertebrae in lizards.23
The objectives of this study were to develop an ultrasound tech-
nique and to describe ultrasonographic anatomy of healthy leopard
geckos including identification of the organs that are normally visible
on ultrasound. Based on published studies in others lizards,3–6 we
could expect to be able to visualize and describe numerous coelomic
organs such as the liver, the digestive tract (which is simple and
divided in three main parts: longitudinal stomach, small intestines,
and colon),24 the reproductive tract, and the fat bodies of our leopard
geckos. Given the small size of this species, we expected an impaired
visualization of very fine structures such as adrenal glands, pancreas,
or spleen.
2 MATERIAL AND METHODS
This prospective and descriptive study combined both anatomic and
analytical study designs. This research project was approved by the
Institutional Animal Care and Use Committee (IACUC) of the Univer-
sité de Montréal. Eleven subadult or adult female leopard geckos were
acquired from a breeder. For inclusion, all animals were required to
have normal physical examination, have a complete tail, have no his-
tory of laying eggs for two months prior to ultrasound acquisition, have
hematological and biochemical parameters within normal limits, and
have fecal examination results that were not considered to be clini-
cally significant. The decisions for subject inclusion were made by two
authors (C.G. and O.C.), who practice exclusively in zoological medicine,
one of them being a Diplomate of the American College of Zoologi-
cal Medicine and the other in training for specialization. The sample
size was determined based on previously published imaging studies in
reptiles.3 Age ranged from 6 to 21 months with all individuals being
of similar size (weight ranged from 33 to 60 g; length ranged from
15 to 20 cm, and width from 2 to 3 cm). The study was performed
during the reproductive season of the species, between February and
April 2016. All leopard geckos received fenbendazole (Panacur, Merck
Animal Health, Québec, Canada) 50 mg/kg orally once at the begin-
ning of the study to reduce parasitic load. All lizards were fed crickets
(Acheta domestica) and mealworms (Tenebrio molitor) from a commer-
cial source and were supplemented in calcium via gut loading. Geckos
had access to water ad libitum. All individuals were housed in three
large terraria (80–120 × 32–53 × 44–60 cm) placed in a temperature-
controlled room. The temperature was set at 23◦ C (73◦ F) and addi-
tional hot spots were provided with heating lamps and heating mats
to obtain an optimal temperature gradient for the species ranging from
28 to 30◦ C (82–86◦ F) during daytime and 22 to 24◦ C (71–75◦ F) during
nighttime. Paper was used as substrate and several hiding boxes were
provided. Ultraviolet-B light sources were available as advised for this
species.25 The photoperiod was set at 12:12 light:dark cycle.
F I G U R E 1 Photograph showing the position of a leopard gecko
for coelomic ultrasound [Color figure can be viewed at wileyonlineli-
brary.com]
Venipuncture were performed under isoflurane light anesthesia
with an insulin syringe either in the ventral coccygeal vein using a lat-
eral approach or in the cranial vena cava as needed to avoid auto-
tomy, whenever geckos displayed tail movements. Complete blood cell
count was performed in all individuals using Diff-Quick stained smears.
A white blood cell count estimate was obtained by manual cell count
over 10 microscope fields. White blood cell differential was used to cal-
culate absolute leukocyte values based on corrected complete white
blood cell estimate. Due to limited sample volume, plasma biochem-
istry panels were obtained for nine of 11 geckos from a reference lab-
oratory (Idexx Reference Laboratories Ltd., Markham, ON, Canada).
The following biochemical parameters were measured: uric acid, phos-
phorous, calcium, total protein, aspartate aminotransferase, creatine
kinase, and bile acids.
Ultrasound examinations were performed in one session by a sin-
gle board-certified veterinary radiologist (I.M.). Geckos were fasted for
24 h prior to ultrasonographic examination. Geckos were sedated with
alfaxolone (Alfaxan, Jurox, New South Wales, Australia) 5 to 15 mg/kg
in the triceps muscle, as previously described.26 Animals were manu-
ally restrained in dorsal recumbency for coelomic organ and tail exam-
inations (Figure 1), and in ventral recumbency for head examination.
Mean ultrasonography duration was 24 ± 9.2 min (range 8–32 min).
Ultrasound examinations were performed with a 13 to 18 MHz linear
array transducer using an ultrasound unit (Toshiba Aplio 400, Toshiba
Medical, Markham, ON, Canada). Gain, focal zone, and time-gain com-
pensation were kept constant for all ultrasonographic examinations.
Ultrasound aqueous gel was applied on the skin. Sagittal, transverse
images, and video captures were acquired by the radiologist (I.M.) and
stored. Animals were immediately recovered and returned to their
normal housing. At a later time, the stored images were analyzed. Visi-
bility, size, localization, and echogenicity were recorded for each organ.
Length, height, and/or width were recorded depending on the organ.
COJEAN ET AL . 3

Length was determined as the distance between the most cranial and
the most caudal parts of an organ; height was defined as the distance
measured from the most dorsal to the most ventral aspects. Width
was defined as the distance measured from the right lateral to the left
lateral margins of the body. Measurements were performed by two
authors (O.C. and I.M.) with electronic calipers, using a DICOM images
viewer (Horos 64 bit v. 2.0.2, Annapolis, Maryland, USA) and a Java
image processing software (ImageJ, Madison, USA), on a personal com-
puter (MacBook Pro, OS X, Apple). The recorded data were the maxi-
mal dimensions obtained for each organ. Liver, ovarian follicles, and fat
bodies echogenicity were compared to each other. Since these three
organs were not visualized on the same image, echogenicities were
compared two by two: those of the liver and the ovarian follicles and
those of the ovarian follicles and the fat bodies.
Six of the 11 leopard geckos were euthanized 1 week following
ultrasound examinations and underwent complete necropsies. Briefly,
geckos were anesthetized with isoflurane in an induction chamber
and administered an overdose of intravenous barbiturate (Euthansol,
Merck Animal Health, Québec, Canada). Necropsies were performed
by two authors (C.G. and O.C.). These two authors were aware of ultra-
sound findings. Samples of the following organs were fixed in forma-
lin and submitted to the pathologist: skin, oral mucosa, tongue, eye,
ear, brain, trachea, lung, heart, digestive tract, liver, reproductive tract,
kidney, and fat pad. Formalin fixed tissues were processed routinely,
embedded in paraffin, sectioned at 5 𝜇m, and stained with hematoxylin,
F I G U R E 2 Schematic drawing of anatomical topography of coelomic
eosin, and safran.
organs of the leopard gecko. The leopard gecko's right is to the left
Descriptive statistics (median, standard deviation, minimum, and side of the image. (Copyright 2016 by Delphine Grosset, reprinted with
maximum) were performed by one author (C.G., Diplomate of the permission) [Color figure can be viewed at wileyonlinelibrary.com]
American College of Zoological Medicine with prior experience in sta-
tistical analyses); and calculated using Excel software (Microsoft Cor-
poration, Redmond, Washington, USA). The ventral surface of the lungs was the most cranially observed
coelomic structure in all animals (n = 10) and the heart was not well
visualized. Lungs were localized cranially and dorsally to the liver and
3 RESULTS their caudal portion was adjacent to the dorsal surface of the liver
(Figures 2 and 5). On transverse images, lungs were observed cranially
None of the leopard geckos had to be excluded from the study due to on the right and the left sides. The ventral pulmonary surface was
abnormal findings on any of the tests. A moderate number of oxyurid characterized by a hyperechoic tissue–gas interface. Deep to this
eggs were detected. No adverse effect was observed during and after hyperechoic line, reverberation or comet-tail artifacts were observed
coelomic ultrasound examinations. in all individuals (Figure 5).
A schematic drawing of the topographic anatomy of the coelomic The liver was easily identified in the second third of the cavity as
organs of leopard gecko was made on the basis of necropsy findings the largest coelomic organ in all individuals. The liver was observed
(Figure 2). The measurements obtained for each organ visible and mea- caudally and ventrally to the lungs, and cranially to the digestive tract,
surable by ultrasound are included in Table 1. the follicles, and the fat bodies (Figure 2). The ventral surface of the
One individual was molting on the day of ultrasonographic examina- liver bordered the ventral coelomic wall. The hepatic parenchyma was
tion. This manifestation precluded visualization of any structure in this homogeneous and its echogenicity was subjectively deemed similar to
individual as ecdysis caused an artifact resulting in diffuse distal acous- that of mammalian liver (Figures 5–7). The liver was hypoechoic to
tic shadowing on ultrasound images (Figure 3). Thereby, the following the follicles and the fat bodies in nine geckos. In one individual, the
structures were visualized in 10 of 11 individuals: ventral surface of liver was hyperechoic to the follicles and hypoechoic to the fat bod-
lungs, liver, caudal vena cava, portal vein, gallbladder, ventral abdom- ies. Contrary to the portal vessels with hyperechoic borders, hypoe-
inal vein, aorta, ovarian follicles, fat bodies, tail, and brain. choic hepatic veins, and arteries were not differentiated from hep-
It was possible to visualize the brain in all 10 remaining geckos. On atic parenchyma. Although the number of liver lobes is difficult to
transverse images, the two cerebral hemispheres were symmetric in assess on ultrasound, gross necropsy showed that there were two liver
size and the nervous tissue was homogeneous and overall hypoechoic lobes, respectively the right hepatic lobe and the left hepatic lobe. On
compared to the surrounding structures (Figure 4). gross necropsy, all animals had a discolored liver. Histopathological
4 COJEAN ET AL .

TA B L E 1 Measurementsa obtained for organs or structures visible and measurable by ultrasound in leopard gecko (Eublepharis macularius)
(n = 10)b

Median SDc Minimum Maximum

Brain Height 7.23 1.53 4.81 8.94
Width 10.41 1.41 8.59 13.68
Liver Height 9.36 1.84 6.15 12.16
Width 19.76 3.34 14.37 24.10
Caudal vena cava (intrahepatic portion) Height 1.49 0.36 0.63 1.78
Portal vein Height 1.04 0.21 0.93 1.49
Ventral abdominal vein Height 0.99 0.17 0.68 1.14
Aorta Height 1.09 0.23 0.80 1.45
Gallbladder Long-axis diameter 3.31 0.80 2.58 4.87
Short-axis diameter 3.47 0.49 2.81 4.43
Follicles Height 9.01 1.54 6.53 11.47
Width 10.90 3.17 5.57 16.75
Fat bodies Length 17.16 2.34 12.77 19.77
Height 6.31 1.35 4.48 8.35
Width 12.52 2.33 11.02 18.33
Tail Height 11.61 1.59 7.95 13.18
Width 15.03 2.61 10.37 18.39

Notes. n, number of individuals; SD, standard deviation.

a Measurements in mm.
b n, number of individuals.
c SD, Standard deviation.

F I G U R E 3 Sagittal ultrasonographic image of the coelom of a molting leopard gecko. Notice the artifact (up arrow) resulting in diffuse distal
acoustic shadowing, precluding visualization of any internal structure. Cranial is to the left and ventral is to the top
COJEAN ET AL . 5

F I G U R E 4 Transverse ultrasonographic image of the brain in a leopard gecko.53–55 Note the gel layer above the skull, necessary to obtain this
ultrasonographic image. The leopard gecko's right is to the left side of the image and dorsal is to the top

F I G U R E 5 Sagittal ultrasonographic image displaying the location of the liver ventral to the lungs in a leopard gecko. Notice the hyperechoic
tissue–gas interface of the lungs (down arrow) and the reverberation artefact below. Cranial is to the left and ventral is to the top
6 COJEAN ET AL .

F I G U R E 6 Sagittal ultrasonographic image showing the position of the liver cranial to an ovarian follicle (up arrow) in a leopard gecko. Notice the
intrahepatic portion of the portal vein (down arrow). Cranial is to the left and ventral is to the top

F I G U R E 7 Transverse ultrasonographic image of the liver in a leopard gecko. Notice the well circumscribed circular anechoic gallbladder, located
near the ventral wall and to the right of midline (up arrow). The leopard gecko's right is to the left side of the image and ventral is to the top
COJEAN ET AL .
analysis revealed hepatocellular vacuolization, characterized by the
presence in hepatocytes of multiple empty vacuoles of variable sizes,
up to twice the nucleus diameter, and filling two-thirds of the entire
cytoplasm. This change was diffused and homogeneous throughout the
hepatic parenchyma and was considered physiologic for reproductive
females.
The caudal vena cava was visualized in all animals at the cranioven-
tral aspect of the liver, on the right side. The caudal vena cava emerged
from the liver cranially to reach the heart. The caudal vena cava was
the largest vascular structure identified in the present study.
The portal vein was easily observed in all animals on sagittal images
on the medioventral aspect of the liver, caudally to the gallbladder.
It was possible to follow the portal vein entering the liver caudally
(Figure 6).
The ventral abdominal vein was visualized in all individuals, extend-
ing along the ventral wall of the coelom, from the caudal aspect of the
fat bodies to the caudal aspect of the liver. It was possible to follow
the ventral abdominal vein over a variable length depending on the
individual.
The aorta was visualized in all individuals, immediately ventral to
the vertebrae. It was possible to follow the aorta from the caudal
aspect of the pleural surface to the level of the fat bodies.
The wall of all described vessels was hyperechoic while the lumen
was anechoic.
The gallbladder was visualized in all individuals on sagittal and
transverse images and was located at the ventral aspect of the liver,
slightly to the right of midline (Figure 2). The gallbladder was oval with
smooth and regular margins (Figure 7). The gallbladder content was
anechoic in all 10 animals and its wall was extremely thin, below the
precision of the calipers, and thus, unmeasurable. Other biliary struc-
tures were not identified in any animal.
The digestive tract was visualized in all animals, dorsally and cau-
dally to the liver, in the medial and the caudal thirds of the coelom. Cau-
dal portions of the digestive tract were visualized medially between the
fat bodies on transverse images (Figure 2). Intestinal wall layering was
not discernible and diameters of visible digestive segments were not
measurable due to resolution limitations. The different segments of
the digestive tract could not be individually identified on ultrasounds.
Some segments contained dietary content while others were empty.
The spleen, pancreas, and adrenal glands were not identified in any
animals.
Urinary structures, particularly the kidneys and urinary bladder
could not be identified in any animal by either ventral or dorsal
approaches.
The reproductive tract was easily identified due to the development
of ovarian follicles. Ovarian follicles were visualized bilaterally in eight
out of the 10 animals and unilaterally, on the right, in one individual.
Two to three follicles were counted on sagittal and transverse images
depending on the individual. Follicles occupied a large portion of the
dorsal coelom and were bounded cranially by the caudal aspect of
the liver and caudally by the cranial aspect of the fat bodies (Figure
2). Follicles echogenicity was lower than that of fat bodies and higher
than the liver echogenicity in eight out of nine animals. In one indi-
vidual, follicles were hypoechoic to both the liver and the fat bodies.
7
Follicles echogenicity was homogeneous in five out of nine geckos and
heterogeneous in four out of nine geckos. One follicle in one of these
four individuals presented a central anechoic area and a hypoechoic
periphery. Follicles were rounded with smooth and regular margins
(Figure 6). Other reproductive structures could not be identified in any
animals.
Multiple rounded, vascularized, and turgid ovarian follicles were
present in all necropsied individuals. While we counted two to three
follicles in each leopard gecko on ultrasound images, up to four folli-
cles were present on necropsy in one individual. The individual who did
not show ovarian follicles did not undergo necropsy. Vitellogenic fol-
licles were described on histopathology reports in all individuals and
a large number of acidophilic granules in the uterine glandular cells
were observed in four out of six individuals undergoing postmortem
examination.
Fat bodies were visualized in all ten nonmolting leopard geckos,
consisting of the second largest coelomic organ after the liver. This
paired structure was located ventrally in the most caudal part of the
coelom, bounded cranially by the liver, and ventrally by the coelomic
wall (Figure 2). On transverse images, these structures appeared bilat-
erally and were symmetric (Figure 8). Fat bodies echogenicity was
homogeneous. These adipose structures were more hyperechoic, com-
pared to the liver and follicles in all animals.
Ultrasound examination of the tail was performed on each leopard
gecko. On transverse images, the spinous process, transverse process,
and body of the hyperechoic coccygeal vertebra were identified, at the
level of the largest diameter of the tail.
4 DISCUSSION
Ultrasound examination with a 13–18 MHz linear array transducer
allowed the visualization and measurement of the following coelomic
structures in this sample of healthy, female leopard geckos: ventral
surface of lungs, liver, caudal vena cava, portal vein, gallbladder, ven-
tral abdominal vein, aorta, ovarian follicles, and fat bodies. Necropsies
were conducted in several individuals to provide comparison between
ultrasound images and postmortem examinations. The lack of normal
references in this species, prompted us to compare our morphological
sonographic descriptions and measurements to those of other lizard
species.3–6,27,28 Some of the organs described in larger lizard species
were not detected on ultrasound in leopard geckos of our study. We
believe that the lack of detection of certain structures was due to their
small size and resolution limitations of the imaging equipment. Since
the weight of a green iguana is about 10 to 30 times that of a leop-
ard gecko and that the weight of a bearded dragon is about five to 10
times that of a leopard gecko, it is possible that the small size of some of
the coelomic organs prevented us from obtaining reliable measures or
visualizing certain organs in leopard geckos. High frequency transduc-
ers offer an enhanced image quality. In this study, the available ultra-
sound unit was the one commonly used for dog and cat imaging by the
Radiology Service. It is likely that higher frequency (20–25 MHz) trans-
ducers with ultrasound unit geared toward use in small animals such as
rodents would provide better images in leopard geckos.
8 COJEAN ET AL .
F I G U R E 8 Transverse ultrasonographic image of the symmetric left and right fat bodies in a leopard gecko. Notice the caudal portions of the
digestive tract located medially (up arrow), between the fat bodies. The leopard gecko's right is to the left side of the image and ventral is to the top
Ultrasonography has numerous advantages in animal species
including lizards. It is a noninvasive imaging modality that does not
require anesthesia. Depending on the temperament of the animal, a
manual restraint or a slight sedation can be used. This is always pre-
ferred, given the challenge of performing safe anesthesia in reptiles,
because of their unique anatomy and physiology. Anesthetized reptiles
commonly present respiratory depression, characterized by bradycar-
dia or apnea and need manually or mechanical assisted ventilation.29
As other small reptile species, leopard geckos are susceptible to
hypothermia and supplemental heat should be provided during anes-
thesia. Recovery can also be long in reptiles and lizards need to be
monitored closely. Detailed description of normal sonographic appear-
ance of coelomic organs in leopard geckos is essential to reliably dis-
criminate abnormal from normal organs and to better characterize the
anomalies using ultrasound. Moreover, it is also relevant to know which
organs are not normally visible by ultrasound. As in mammals, ultra-
sonography provides pertinent clinical information that aid in the diag-
nosis of several gastrointestinal, hepatic, or reproductive diseases.1
Other diagnostic procedures such as ultrasound-guided fine needle
aspirates can be performed1 in exotic animals as in mammals. Further-
more, ultrasonography may be helpful to follow the evolution of the
reproductive cycle in female leopard geckos. It could provide a tool
to monitor follicular development and to determine the presence or
absence of eggs.
Some features of reptile species are responsible for creating arti-
facts in ultrasound. The main artifacts encountered are due to the pres-
ence of air in the lungs creating distal acoustic shadowing and masking
the most dorsal structures (aorta, bones structures); and the presence
of air trapped between irregular skin and scales.30,31 Due to the large
amount of air present between the layers of keratin, ecdysis also
prevented the penetration of ultrasound waves, as observed in one of
the 11 leopard geckos of this study.
Due to the small size of the species studied and the thinness of
their skull, ultrasound of the brain was made possible (Figure 4). With
a standardized method, ultrasound could be an alternative safer and
less expensive imaging modality than magnetic resonance imaging for
the diagnosis of diseases of the central nervous system such as tumor
masses, inflammatory granulomas or hydrocephalus.18 Although these
pathologies have rarely been described in lizards, there are some
reports of cases of cerebral xanthomatosis in geckos15 and green water
dragons.16 It is interesting to develop alternative of magnetic reso-
nance imaging because this modality requires long and closely moni-
tored anesthesia, which is less safe as explained above. Also, although
magnetic resonance imaging is a most powerful device, it could be less
sensitive for very small species as leopard geckos. Finally, ultrasound
requires equipment more accessible for smaller veterinary structures,
which do not have access to magnetic resonance imaging.
The appearance of the ventral surface of lungs, characterized by
a hyperechoic interface with reverberation artifacts below (Figure 5),
was similar to that observed in bearded dragons.3 This appearance is
normal and characterizes the tissue–gas interface. This should not be
confused with ring-down artifacts, which are signs of extensive alveo-
lar or interstitial disease (such as pulmonary edema, pulmonary fibro-
sis, interstitial pneumonia, or pulmonary contusion) in humans and
COJEAN ET AL .
dogs.32 Leopard geckos display unicameral lungs formed of a single
chamber per lung and do not have any air sac,33 hence the ultrasono-
graphic appearance is homogeneous throughout the lung surface.
Although the heart was not visualized in any of the leopard geckos
in this study, a few published references describe the echocardio-
graphic anatomy in lizards,3,4,34,35 chelonians,36 and snakes in the fam-
ily Boidae.37 In bearded dragons3,35 and green iguanas,4,34 the heart
was visualized in the cranial and ventral aspects of the coelom, cranially
to the liver, and cardiac chamber measurements have been reported in
these species.
Although the gastrointestinal tract was observed in all animals,
the different parts of the digestive tract could not be individually
delineated and the diameter of the visible digestive portions was not
measurable in any individual of this study. Conversely in bearded
dragons,3 the fundic and pyloric portions of the stomach, small intes-
tine, and cecum were identified and measured using ultrasonography.
In bearded dragons, the wall of these digestive structures was as thin
and discrete as in leopard geckos, but their thickness could be mea-
sured. Likewise in the green iguanas,4 only the pyloric portion of the
stomach was well differentiated and measured. Perhaps it is possible
that the leopard gecko being a much smaller species than the latter two
could explain the difficulty in obtaining precise measures of the gas-
trointestinal tract in our study.
The spleen was not visualized in any of the leopard geckos of this
study. Although this organ has been described in green iguanas,4 its
visualization was not possible in bearded dragons3 and bosc monitors5
with a 5–15 MHz transducer . The spleen, pancreas, and adrenal glands
may be too small in leopard geckos to resolve them using ultrasound
units, purposely acquired to be used in domestic small animal species
such as dogs and cats.
The urinary tract was not identified in any individual with either
dorsal or ventral approach. In leopard geckos, the paired kidneys are
the most caudal coelomic structures, located within the pelvis.38 Their
small size and location, being surrounded by bony structures, may have
prevented us from their visualization. Ultrasonographic images of kid-
neys are available in large lizard species including bearded dragons,3
green iguanas,4 and bosc monitors.5 In bearded dragons, a ventral
approach in the transverse plane allows the identification of the cra-
nial poles of the kidneys and a dorsal approach facilitates distinction of
the cranial and caudal poles of each kidney. In green iguanas, the cra-
nial pole of the kidneys were only seen in four of the 26 animals.4 Due
to their large size, the ultrasonography of kidneys’ bosc monitors was
also possible and described.5
Although the presence of a urinary bladder has been reported in
green iguanas, geckos,39 and chameleons among others,38,40 ultra-
sonography of this structure is only described in green iguanas among
lizards.4 Since the bladder is sometimes rudimentary and has a very
thin wall, the urinary bladder was not always seen in gross necropsy of
leopard geckos, and may explain the difficulty in identifying this organ
on ultrasound.
In our leopard geckos, the ultrasonographic appearance of the liver
was subjectively similar to that of various species, including dogs and
cats. The hepatic parenchyma was homogeneous and the liver had
sharp and regular edges. The only discernable hepatic vessels were the
9
portal vessels, which were characterized by hyperechoic walls similar
to that described in mammals liver anatomy.41 When imaged in a trans-
verse plane, the liver of leopard geckos was subjectively divided in two
lobes of similar size separated on midline. This assumption, similar to
previous data,42 was confirmed at necropsy. Despite severe hepatocel-
lular vacuolization observed histologically in all necropsied geckos, the
liver remained hypoechoic to the fat bodies in all individuals preventing
the ultrasonographer from making a presumptive diagnosis of hepatic
lipidosis. The fact that biochemistry results, including aspartate amino-
transferase enzyme, bile acids, and total proteins, were within normal
limits also suggests that hepatocellular vacuolization was physiologic
in these reproductive female leopard geckos.
Since adipose tissue is among the most hyperechoic tissues of the
coelomic cavity, fat bodies of lizard species may offer a useful base-
line tool for the evaluation of various structure echogenicity in the
coelom.1 As expected,5,8,17,19 the echogenicity of the fat bodies was
superior to that of the follicles and the liver in ten individuals. As in
mammals, hepatic lipidosis in reptiles is characterized by an increase
in echogenicity of the liver and less discernable vascular structures in
the hepatic parenchyma.3,18 It is often not possible to differentiate the
liver and fat body from each other during an ultrasound examination
when a reptile is suffering from hepatic lipidosis.1 In extreme cases,
the margins of the liver are rounded due to an increased organ size.1
Hepatic lipidosis is pathologic and should be differentiated from physi-
ological hepatocellular vacuolization occurring during vitellogenesis in
female lizards.43 This is a physiological mechanism occurring in female
reptiles due to buildup of hepatic lipid stores43 in anticipation of the
reproductive period.
Females included in this study were in the vitellogenic phase of their
reproductive cycle. Leopard geckos have a seasonal monoautochronic
ovulation pattern,44 defined as both ovaries typically ovulating one
follicle synchronously and females laying clutches consisting of two
eggs most commonly, although single-egg and triple-egg clutches are
occasionally encountered.45 Due to the large size of the ovaries, it
was not always possible to lateralize the follicles. Moreover, there
was some discrepancy between the numbers of follicles identified
on ultrasound compared to necropsy, the former tending to under-
estimate the total number of follicles. An accurate estimation of the
number of follicles is difficult on ultrasound since these structures can
be superimposed over each other and not individually delineated or
the same follicle can be counted several times. In addition, necropsy
was not performed immediately after ultrasound examination, and
ovarian development could have progressed in the interim. Follicles
appeared diffusively homogeneous or heterogeneous depending on
individuals. In veiled chameleons,6 other lizards species,46,47 and
turtles,48,49 ultrasonography is a reliable method to describe various
stages of follicles and to distinguish them from eggs. Based on previous
studies in lizards, the comparison between follicles and eggs could
rely on their morphological appearance: follicles are round structures,
while eggs are ovoid with a hyperechoic outline, corresponding to the
shell.27,50–52 Also, according to Schumacher and Toal, preovulatory
follicles are hypoechoic, whereas postovulatory follicles are more
hyperechoic.50 In the present study, no egg was observed by ultra-
sound, or at postmortem examinations. Moreover, ultrasonography
10
can help differentiation between new and old batches of follicles.6
Postmortem ultrasonography of a veiled chameleon ovary indicated
that newer follicles display hyperechoic concentric rings throughout
the follicular structure, whereas older follicles have a hypoechoic
center.6 This later feature was observed only in one individual of
the present study. Whether a hypoechoic follicular center is asso-
ciated with older follicles in leopard geckos would warrant further
studies.
A limitation of the present study was the use of only repro-
ductive females during the vitellogenesis phase of their cycle. We
chose this population to describe the normal ultrasonographic appear-
ance of vitellogenic follicles. Additional studies would be needed
to describe the female reproductive system outside of the repro-
ductive period. Furthermore, a longitudinal study monitoring the
sonographic evolution of the reproductive system of female leopard
geckos and assessing potential associations between these morpho-
logic changes during the different stages of their cycle may be of
benefit in this species. In addition, it would be interesting to per-
form ultrasonographic examination of male leopard geckos. We know
that the reproductive system of both male bearded dragons3 and
green iguanas4 can be observed by ultrasonography, in particular their
testis and hemipenes. Ultrasonographic imaging of normal male repro-
ductive organs could then facilitate the identification and diagnosis
of pathologies of the male reproductive system in individuals pre-
sented to the clinic. Finally, our results were limited by the small
size of the species studied and the resolution provided by the high-
est frequency linear transducer (13–18 MHz) available in our radiol-
ogy service, which did not allow the visualization of certain coelomic
organs. However, leopard geckos are frequently encountered at our
clinic of zoological medicine and ultrasonography represents an afford-
able noninvasive imaging modality to complement other diagnostic
procedures.
In conclusion, results of this study supported the use of ultrasonog-
raphy as a diagnostic tool for the assessment of coelomic and non-
coelomic structures in leopard geckos. Ultrasound examination with
a 13–18 MHz linear array transducer allowed the visualization and
measurement of the following coelomic structures in leopard geckos:
ventral surface of lungs, liver, caudal vena cava, portal vein, gallbladder,
ventral abdominal vein, aorta, ovarian follicles, and fat bodies. The
digestive tract was visualized but the different digestive segments
could not be individually identified and measured. Ultrasonography
did not allow the visualization of the spleen, pancreas, adrenal glands,
and urinary structures in any animal. Further studies would be needed
to describe the ultrasonographic anatomy of the male leopard gecko.
LIST OF AUTHOR CONTRIBUTIONS
Category 1
(a) Conception and Design: Cojean O, Vergneau-Grosset C, and
Masseau I
(b) Acquisition of Data: Cojean O, Vergneau-Grosset C, and
Masseau I
COJEAN ET AL .
(c) Analysis and Interpretation of Data: Cojean O, Vergneau-
Grosset C, and Masseau I
Category 2
(a) Drafting the Article: Cojean O and Vergneau-Grosset C
(b) Revising Article for Intellectual Content: Cojean O,
Vergneau-Grosset C, and Masseau I
Category 3
(a) Final Approval of the Completed Article: Cojean O,
Vergneau-Grosset C, and Masseau I
ACKNOWLEDGMENTS
The authors would like to acknowledge Dr. Stéphane Lair for
histopathology results, the technician team of the Diagnostic Imag-
ing Service, and of the Centre Québécois sur la Santé des Animaux
Sauvages of the Université de Montréal for their assistance during
ultrasounds and tissue processing, respectively, and the Association of
Reptilian and Amphibian Veterinarians for providing funding for part
of this project.
ORCID
Ophélie Cojean http://orcid.org/0000-0003-1308-9203
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How to cite this article: Cojean O, Vergneau-Grosset C,
Masseau I. Ultrasonographic anatomy of reproductive female
leopard geckos (Eublepharis macularius). Vet Radiol Ultrasound.
2018;1–12. https://doi.org/10.1111/vru.12599