Vet Clin Exot Anim 7 (2004) 673695

Amphibian oncology
Brian A. Stacy, DVMa,*, John M. Parker, DVMb
a

Anatomic Pathology Service, Veterinary Medical Teaching Hospital,

University of CaliforniaDavis, One Shields Avenue, Davis, CA 95616, USA
b
Oce of Laboratory Animal Care, Northwest Animal Facility,
University of CaliforniaBerkeley, Room 203C, Berkeley, CA 94720-7150, USA

The class Amphibia consists of three orders: Anura (frogs and toads),
Urodela (a.k.a. Caudata) (salamanders), and Gymnophiona (caecilians).
Captive and wild populations are important to many areas of scientic
research, and are a focus of major global ecologic concerns, including
pollution, climate changes, habitat destruction, and nonnative species translocation. Such concerns have been identied as causes of decline in certain
amphibian populations. Also, the number and diversity of wild and captive
amphibians maintained and monitored by zoologic and teaching institutions,
research facilities, and hobbyists continues to grow. Consequently, veterinarians are consulted more frequently for information on health and disease.
Reports of spontaneous neoplasia in wild amphibians are relatively rare
and often limited to specic species or populations [1]. For example, viralinduced Lucke adenocarcinoma is documented only in northern leopard
frogs (Rana pipiens), and a high incidence of neoplasia in barred tiger
salamanders (Ambystoma mavortium) is limited to a population inhabiting
a polluted pond in Texas. Research has shown that at least some anuran
species possess inherent anticancer secretory products and cytoprotective
devices, and are relatively resistant to some mammalian carcinogens [27].
Also, the remarkable regenerative capacity of urodeles is hypothesized to
reduce tumor susceptibility [8,9]. To date, neoplasia has not been reported in
Gymnophiona.
Amphibians are comprised of the same basic tissue elements as other
species, and any cell type can undergo neoplastic (malignant) transformation. The biologic behavior of a given tumor type may vary among
amphibian species because of natural species variation and etiologic
dierences, such as viral infection, exposure to pollutants, or inheritable

* Corresponding author.
E-mail address: bastacydvm@glasselevator.net (B.A. Stacy).
1094-9194/04/$ - see front matter 2004 Elsevier Inc. All rights reserved.
doi:10.1016/j.cvex.2004.04.001

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(genetic) predisposition [1013]. Clinical management is facilitated by

understanding documented features of tumors, which may provide insight
into treatment options, identication of predisposing factors, and management of populations. Furthermore, denitive diagnosis is important because
tumors with an infectious or acquired etiology and infectious pseudoneoplastic conditions, such as mycobacteriosis, are relatively common, and
may spread within the immediate group or population.
Literature pertaining to amphibian neoplasia is scattered, often incomplete, and dicult to interpret due to inconsistencies in diagnosis, nomenclature, and misdiagnosis of infectious and inammatory conditions.
Furthermore, clinically relevant information, such as long-term prognosis,
biologic behavior, and therapy, is lacking, and many reports focus on aspects
of tumor research rather than clinical management. Green and Harshbarger
[14] provided a recent and thorough review of published cases and those
submitted to the Registry of Tumors in Lower Animals (RTLA) (22,900 Shaw
Road, Suite 107, Sterling, Virginia 20166-4311, USA). Such references, as well
as materials maintained by the RTLA, are useful resources for veterinarians.
This article focuses on relatively common types of neoplasia, methods for
antemortem diagnosis, dierential diagnoses, and techniques for surgical
excision. Select cases from the large amphibian population maintained by
the University of California, Berkeley (UCB) are used to exemplify some
tumor types. Classically, the terms tumor and neoplasia have dierent
denitions, and their use in the literature has led to some confusion. Most
current references, however, use these terms interchangeably and regard
them as synonyms. Thus, both terms are used interchangeably throughout
this text. A variety of tumors have been induced experimentally, and those
without clinical relevance are not considered here.

Tumors of the integument and soft tissues

Neoplastic diseases of the integument are documented in several anuran
and urodele species, and constitute a large proportion of the literature on
amphibian neoplasia [14,15]. Cases include neoplasia of the surface and
glandular epithelium, dermal stroma, and melanophores. In some instances,
tumors have unique features such as environmental and infectious etiology,
temperature-dependent growth, and predictable regression under laboratory
conditions. Many neoplasms are only reported in specic populations, and
their occurrence and biologic behavior in other wild and captive populations
is unknown.
Epidermal papillomas
Epidermal papillomas, also referred to as squamous papillomas, infectious warts, and epitheliomas, are described most frequently in urodele

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species, including free-ranging Japanese newts (Cynops pyrrhogaster),

barred tiger salamanders (A mavortium) and Tohoku salamanders (Hynobius lichenatus) [11,1619]. These papillomas are sessile, hypopigmented,
and well demarcated with a folded or convoluted surface [16,17]. Histologically, they consist of marked epidermal hyperplasia that forms papillary
projections and occasionally involves the glandular epithelium and extends
into the dermis [17,18]. Most are solitary lesions of little health concern, and
are readily debulked or excised for diagnosis. In a few cases, however,
papillomas were multicentric, and associated with poor body condition
[17,18].
Spontaneously regressing skin papillomas in wild C pyrrhogaster occur
seasonally. The highest prevalence is in autumn, with aected newts
comprising up to 5.4% of the animals examined [20]. In a case series of
10, seven regressed over 4 to 12 months and three progressed to large,
multicentric lesions [18]. In another series, tumors in 44 newts regressed over
a period of months when placed under laboratory conditions [17]. Studies
have shown that temperature strongly inuences tumor growth and
regression. Tumors of newts housed at 4(C and 25(C regress in size and
even sloughed o in some cases, and tumors increase in size at midrange
temperatures of 10(C to 13(C [21]. Also, experimental exposure to
ultraviolet radiation caused regression in one report [22]. A viral etiology
is suspected based on the detection of herpesvirus-like particles in some
tumors [20,23]. This suspicion was not pursued further, and tumor transmission could not be induced experimentally [17].
Regressing papillomas are also documented in a neotenic population of A
mavortium that inhabits a polluted sewage pond [11]. These tumors are
presumably caused by an environmental stimulus, having regressed after 4
months under laboratory conditions in a controlled study [11]. Whether
papillomas will eventually regress spontaneously in most other species is not
known.
In anurans, reports of papillomas are limited to a single case in a black
spotted frog (Rana nigromaculata) and a case series in R pipiens [14,24,25].
In addition, two long-term captive Surinam toads (Pipa pipa) housed at the
UCB, developed large, insidious, pedunculated papillomas on the distal
extremities (Fig. 1), both of which were excised successfully. This species is
known for having warts, and an underlying viral etiology is suspected.
Despite prolonged cohabitation, however, evidence for high infectivity was
not observed in these cases [25; JMP, personal observation].
Other epithelial tumors
Other epithelial tumors reported in amphibians include squamous cell
carcinomas and dermal gland tumors. Van der Steen et al [24] examined a series
of cutaneous tumors in R pipiens that included 10 squamous cell carcinomas.
These tumors were characterized grossly as hypopigmented, raised, sessile, or

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Fig. 1. A pedunculated papilloma extends from the dorsal aspect of the distal digit of a Surinam
toad (Pipa pipa). The tumor was easily excised by ligation of the brovascular stalk.

pedunculated, 0.3 to 1.5 cm diameter masses. Histologically, tumors were

comprised of inltrative nests of nonkeratinizing epithelium. Most tumors
were locally inltrative but did not invade deep to the dermis, except for one
case involving the deep skeletal muscle and bone. Adenomas and adenocarcinomas of the dermal glands are documented as single case reports, as
well as a larger series of 78 neoplasms in seven grass frogs (Rana temporaria)
and 16 pond frogs (Rana ridibunda) [15]. Representing the majority of the
literature on dermal gland tumors, this series described cystadenomas and
cystadenocarcinomas as comprised of dilated glands lined by papillary
projections of neoplastic epithelium. The diagnosis of cystadenocarcinoma
was based on increased cellular pleomorphism and atypia. Tumors were
conned to the dermis, and metastasis occurred in a single case. Other
tumors of dermal glands include rarely reported mixed epithelial and
myoepithelial (contractile glandular tissue) tumors and a presumed myoepithelial adenoma [14]. Based on these reports, surgical removal of squamous
cell carcinomas and dermal gland tumors is advised and both may exhibit
local inltration and metastasis.
Melanophoromas
Melanophoromas, also referred to as melanoma, melanocytoma, and
melanosarcoma, are described in multiple species of urodeles and anurans.
Neoplasms vary greatly in behavior and gross morphology, ranging from
solitary to multicentric, hypopigmented to pigmented, nonpalpable to
nodular, and regressive to invasive masses. Histologically, as with melanocytic tumors in mammals, they can exhibit several cell types, including
dendritic, epithelioid, and spindle-shaped, but often are classied by the
dominant cell type [14]. In some cases, melanophoromas were limited to the
dermis and did not invade the underlying lymph sac, making them amenable

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to surgical excision [12,26]. Some of these benign forms are analogous to

melanocytoma or melanocytic nevi in mammals. A few highly invasive or
malignant melanophoromas are documented, and include darkly pigmented
and hypopigmented variants [14].
An important dierential diagnosis for melanophoroma is melanophore
hyperplasia secondary to skin diseases (Fig. 2A). Some melanophoromas
initially present as at, darkly pigmented foci and grossly are indiscernible
from hyperplastic lesions; therefore, biopsy is required for denitive
diagnosis [26,27].
Melanophoromas have unique features in some species, such as hereditary
predisposition or possible viral etiology [13,26]. In studies with transplantable melanophoromas in crested newts (Triturus cristatus carnifex), tumor

Fig. 2. (A) An unidentied Xenopus species with focal melanophore hyperplasia (arrow)
associated with dermatitis. The inammation in this case was due to nematodiasis of the dermal
glands and required histopathology for diagnosis. Early melanomas may begin as grossly
similar lesions. (B) The dorsum of a barred tiger salamander (Ambystoma mavortium) with
a large melanophoroma. Melanophoromas in this population regressed under laboratory
conditions and were presumed to have an environmental stimulus. (Courtesy of F.L. Rose,
PhD, Texas State University.)

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growth rate and metastasis are temperature dependent. Namely, tumors were
relatively static at 4(C, but grew rapidly and metastasized at 30(C [28].
Temperature-dependent growth is shared with Lucke renal adenocarcinomas, epidermal papillomas of Japanese newts, and possibly other amphibian
tumors [21,28,29]. Caution must be exercised, however, in extrapolating
information about biologic behavior between dierent species, as dierent
etiologies and biologic variations could result in dierent behavior.
Several cases of melanophoroma are reported in Mexican axolotls
(Ambystoma mexicanum), but little information on age and survivability
is provided [15,26]. In cases reported by Khudoley and Mizgireuv [15],
melanophoromas in both pigmented and albino A mexicanum consistently
seemed to progress from a at proliferation of melanophores to a nodular
growth. For example, a male axolotl developed a tumor at 2.5 months of age
and developed additional masses 16 months later. In a case series presented by
Sheremetiva-Brunst, and Brunst [26], hereditary predisposition was evidenced
by the high prevalence of tumors in ospring from two individuals with
melanophoromas. The second generation developed tumors at approximately
1 year of age, and these grew faster and larger than those of the parent generation. Histologically, neoplasms were conned to the dermis and comprised
predominantly of dendritic and heavily pigmented cells with anisokaryosis,
anisocytosis, and multinucleated cell formation. In both reports, authors
describe two types of tumor growththose with and those without deep tissue
invasion [15,26]. Metastasis was not reported in any case. In summary, based
on these few cases, melanomas in pigmented and albino axolotls may develop
at a relatively early age, may be locally invasive and multicentric, but are not
known to metastasize, and recurrence may follow surgical excision.
In a population of A mavortium that inhabited a sewage pond, melanophoromas were associated with other hyperplastic and neoplastic lesions of
the integument (Fig. 2B) [30]. The gross appearance and potentially locally
aggressive nature of these tumors were similar to those described in A
mexicanum, but regression of some melanophoromas, including inltrative
tumors, followed the removal of animals from the polluted environment and
metamorphosis (FL Rose, personal communication).
Another series of melanophoromas was reported in the African clawed
frog (Xenopus laevis) [13]. Eight frogs out of 40,000 developed single
melanophoromas on the dorsum, ventrum, and limbs. These tumors diered
from many of those observed in A mexicanum and A mavortium, because
they were hypopigmented and highly invasive. Herpesvirus-like viral
particles were observed within tumors; however, it was not determined if
these tumors actually were caused by an oncogenic virus.
Mesenchymal skin tumors
The classication scheme and terminology of dermal mesenchymal
tumors in amphibians is unclear and in need of further characterization.

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The terms dermal broma, polypoid dermal broma, and bropapilloma

have been used to describe a benign nodular proliferation of dermal
broblasts covered by normal or hyperplastic epidermis arising anywhere
on the skin [14]. Dermal mesenchymal tumors are reported in the same
population of A mavortium, which is mentioned throughout this text as well
as in a few geriatric albino A mexicanum at UCB ([30]; B.A. Stacy and J.M.
Parker, personal observation). Grossly, masses are smooth and hypopigmented (Fig. 3A). Histologically, they can be poorly demarcated, and are
comprised of poorly organized streams of neoplastic broblasts with
potentially marked anisokaryosis and generally rare to absent mitoses
(Fig. 3B). The majority of dermal mesenchymal tumors are benign and
exhibit slow growth, and likely do not represent a signicant health threat.

Fig. 3. (A) Geriatric albino Mexican axolotl (Ambystoma mexicanum) with a bropapilloma of
the rostral maxilla. This lesion exhibited slow growth and was an incidental nding. (B)
Photomicrograph of the bropapilloma in (A). The dermis is expanded by a nodular, wellcircumscribed mass of haphazardly arranged streams of neoplastic broblasts. Some neoplastic
cells have a large, bizarre nucleus (arrow). Mitotic gures are not observed. Hematoxylin and
eosin; 20 (inset 400).

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Fibrosarcomas, which may be locally aggressive, are reported in A

mavortium and a few other species [14,31].

Neuroepithelial tumors
Tumors of neuroepithelium are reported in A mexicanum and an alpine
newt (Triturus alpestris) [32,33]. These tumors presumably originate from
olfactory tissue to form space-occupying masses in the maxilla. Histologically, lobules of elongated or bipolar neoplastic epithelium palisade
around cystic spaces containing coagulated material. There is no mention
of long-term survivability. In one case, the tumor was transplanted to
other adult A mexicanum, and one was alive 3 years after successful
transplantation, which may suggest a prolonged clinical course [32].
Tumor location, however, can interfere with feeding and thus present
a signicant health threat. Surgical excision has not been attempted.
Palliative care is recommended until quality of life declines below acceptable standards.

Mast cell tumors

Reports of mastocytomas, also refered to as mast cell tumors, are limited
to urodeles, including A mexicanum, neotenic A mavortium, and red-spotted
newts (Notophthalmus viridescens) [14,33]. A mexicanum were 10 to 17 years
old, and all of A mavortium originated from a polluted sewage pond.
Tumors developed within the dermis or subcutaneous tissues anywhere on
the body, and appeared grossly as domed, rm, gray-white or red, ulcerated,
less than 3.0-cm diameter masses. Histologically, neoplastic mast cells were
polygonal to spindle-shaped with marked anisocytosis and anisokaryosis in
some tumors. These tumors were locally invasive and regional edema in
some tumors suggested the possible release of histamine. Systemic involvement, rate of tumor growth, and longevity following diagnosis were
not addressed in these studies. Giemsa stain revealed metachromatic
granules in 5% to 25% of the neoplastic cells [14]. Initially, many of these
tumors were misidentied as mesenchymal or neural tumors and later were
reclassied after examination with a Giemsa stain [14]. Thus, a Giemsa stain
should be considered when examining similar dermal tumors. Mast cells
should exfoliate well on ne-needle aspirates or impression smears, which
may aid in antemortem diagnosis. It is not known whether amphibians
develop poorly dierentiated, agranulocytic tumors, which could present
a diagnostic challenge. Surgical treatment depends on the location of the
mass. Tumors restricted to the limbs or tail may be cured by amputation of
aected appendage. Tumors on the trunk of urodeles are technically
challenging because skin defects in these animals are not readily amenable
to closure and healing occurs by contraction and epithelialization.

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Hyperplastic skin conditions

As in mammals, hyperplasia is a nonspecic response of the amphibian
integument to injury and typically subsides after resolution of the underlying cause. Cutaneous hyperplasia may present as distinct skin mass(es),
and is caused by a variety of diseases, including parasitic, fungal, and viral
infections or imbalances in water quality [34,35]. In some cases, hyperplastic
lesions are associated with intraepithelial keratin-lled cysts, and have been
documented in association with various skin tumors [24]. The previously
described population of A mavortium inhabiting a polluted sewage pond in
Texas had papillomas and melanophoromas as well as epithelial hyperplasia, which was proposed as a preneoplastic change [30]. Both the hyperplasia
and neoplasia regressed when the animals were removed from the
environment, suggesting an environmental cause.

Tumors of the hemolymphatic system

The anatomy of hematopoietic tissues in amphibians diers from that in
other groups of animals more commonly encountered by veterinarians.
Even within amphibians, there is signicant anatomic variation among
orders and genera, as well as dierent life stages. To those unfamiliar with
amphibian pathology, the character, abundance, and distribution of
hemopoietic tissue and proliferation in response to a variety of stimuli is
a potential source of confusion and misinterpretation. The best example of
this diculty is the large body of literature pertaining to the misdiagnosis of
mycobacteriosis as lymphoma, often referred to as transmissible lymphosarcomata [36].
Lymphoma
The literature on lymphoma in amphibians often is confusing and
contradictory. Much of this confusion results from the misdiagnosis of
mycobacteriosis as lymphoma; therefore, older papers on lymphoproliferative disease in amphibians should be scrutinized carefully. For reference,
proven cases of lymphoma are characterized best in X laevis and A
mexicanum.
A series of T-cell lymphomas was reported in X laevis [37,38]. These cases
primarily involved the thymus, presenting initially as a unilateral swelling of
the caudal cephalic region (Fig. 4A). Histologically, monomorphic neoplastic lymphocytes form densely cellular sheets and may have a high
mitotic rate. Lymphoma may become generalized to involve multiple organs
and progress to leukemia (Fig. 4B). Thus, a diagnosis may be obtained by
complete blood cell count and examination of a blood smear.
Lymphoma in axolotls is the subject of transplantation and histocompatibility research [39]. For spontaneous lymphoma, such features as

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Fig. 4. (A) African clawed frog (Xenopus laevis) with thymic lymphoma resulting in a large
mass in the lateral cervical region. The tumor was soft and white on section and obliterated the
associated skeletal muscle and lymph sac. (Courtesy of Greg Trimmel, UCB) (B)
Photomicrograph of a transverse process of a cervical vertebrae from the X laevis in (A). The
tumor had progressed to leukemia, as evidenced by eacement of the bone marrow by
a monomorphic population of neoplastic lumphocytes. Neoplastic lymphocytes ll the
vasculature of the medullary cavity (asterisks). Hematoxylin and eosin; 100 (inset 1000).

location and progression of disease are not well documented. Lymphoma in

transplant recipients, however, is noted to progress rapidly to leukemia.
Other round cell tumors
The spontaneous occurrence of other types of round cell tumors is limited
to two cases of granulocytic leukemia in a toad (Bufo sp.) [14]. No
additional information is available on these animals.

Tumors of the hepatobiliary system

Neoplasia of the hepatobiliary system is rare in amphibians. As in other
species, accurate diagnosis of liver tumors and the distinction between

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benign and malignant tumors and tumors verses hyperplastic or regenerative nodules may present a diagnostic challenge in some cases. Studies are
needed to characterize the livers natural response to insults and regenerative capability. Some reports suggest similarities between hepatic lesions
and regenerative capability of mammals and amphibians [14]. For example,
hepatic cirrhosis was documented in wild A mavortium [30]. In other species
and perhaps in amphibians, features that help distinguish nodular regeneration from benign hepatocellular adenoma or hepatoma are the
multifocal distribution of the former and absence of normal portal
structures in the latter. The absence of portal structures was described in
an adenoma in the barking tree frog (Hyla gratiosa) [31]. All reported cases
of hepatocellular carcinoma in amphibians were induced experimentally or
associated with environmental carcinogens [14]. Features of malignancy,
including local invasion, poorly dierentiated neoplastic hepatocytes, and
metastasis, may be present; however, in well-dierentiated carcinomas, the
distinction between benign and malignant can be dicult. Spontaneous
tumors of the biliary system in amphibians are also rare. One case was
reported in a group of Asian pond frogs (Rana spp.) with a high prevalence
of pancreatic carcinoma [40]. This hepatic tumor was diagnosed as a biliary
adenoma, but was not well described.
The most common dierential diagnosis for hepatic masses is mycobacterial granulomas. Fine-needle aspirates or impression smears examined
with acid-fast stains are instrumental in making a diagnosis. Additional
dierentials include other causes of granulomas, biliary cysts, and mesothelial hyperplasia, the latter of which may occur in response to chronic
eusion or coelomitis.

Tumors of the gastrointestinal tract

Neoplasia of the gastrointestinal tract is poorly documented and limited
to a few reported cases, most of which were malignant and diagnosed at
necropsy. Elkan [41] described a gastric adenocarcinoma with metastasis in
X laevis. A small number of intestinal adenocarcinomas are described in
various anuran species, although it is often unclear whether the tumors
arose from the enteric mucosa, pancreas, or another organ [14]. Green and
Harshbarger [14] described an intestinal adenocarcinoma in a marine toad
(Bufo marinus) from the RTLA (Herron, RTLA #1921). It was characterized by a prominent scirrhous or desmoplastic response, which is a common
feature of gastroenteric adenocarcinomas in mammals. A group of R pipiens
that were infected experimentally with Mycobacterium marinum developed
intestinal adenomas and adenocarcinomas that resulted in obstruction,
ascites, and in some cases death [42]. The denitive tissue of origin and true
neoplastic nature are uncertain because these tumors were poorly described.
A few cloacal tumors, including an adenocarcinoma in a Mexican axolotl

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and a squamous cell carcinoma of the colorectal junction in an aged

Oriental re-belly toad (Bombina sp.) are documented in the RTLA [14].
A major dierential diagnosis for masses in the alimentary tract includes
granulomas, especially of mycobacterial and parasitic origin, which can be
serosal, intramural, or submucosal. Furthermore, it is recommended that all
suspected stromal tumors of the alimentary tract are examined with an acidfast stain.

Tumors of the urogenital system

Lucke renal adenocarcinoma
Lucke renal adenocarcinoma of R pipiens is the most well-known
amphibian neoplasm. This tumor is caused by a herpesvirus known as
Ranid herpesvirus-1 or Luckes herpesvirus, which is considered endemic in
wild northern leopard frog populations [29]. The Lucke tumor has been the
focus of much research since it was rst described in 1934, and early
accounts date back as far as 1905 [43]. Conrmed cases of spontaneous
nonviral associated renal neoplasia has not been reported in R pipiens, and
only a few cases have been reported in other species.
The pathogenesis of viral infection and tumorigenesis has several
interesting features. Viral transmission is hypothesized to occur from
infected urine in breeding ponds when frogs emerge from winter hibernation
[43]. Two forms of the tumor are recognized. In the winter or algid phase,
tumors are relatively static in size, and viral replication is active. In the
summer or calid phase, tumor growth and metastasis occur, and detectable
viral replication is minimal or absent. Tumor growth and metastasis are
profoundly greater at higher temperatures. Experimental studies demonstrated that large tumors developed at 28(C, whereas colder temperatures
(7(C) inhibited tumor growth, but supported viral replication [45]. The
prevalence of tumors in wild populations uctuates by year, season, region,
and method of detection. In some populations, greater than 90% prevalence
was reported when evaluated for microscopic tumors by histologic
examination [44].
Leopard frogs are a common model used for research and education.
Recently, their taxonomy was reclassied, which facilitated understanding
of the regional distribution of this neoplasm. Any true R pipiens should be
considered potentially infected with this oncogenic virus. Thus, a signicant
number of wild-caught R pipiens may develop tumors under laboratory
conditions where they are maintained at high ambient temperatures. This
can be disruptive to research as large or metastatic tumors will result in
death or necessitate euthanasia. Frogs develop carcinomas only if exposed
to the virus as eggs or young embryos [4649]. Furthermore, infections in
other anuran species have occurred only under experimental conditions.

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There are currently no major eorts underway to establish virus-free R

pipiens colonies.
Clinically, R pipiens with Luckes tumor may present with coelomic
distension, palpable intracoelomic masses, atrophy of thigh musculature,
pelvic limb paralysis, lethargy, and sudden death. Often, clinical signs are
not apparent until tumors are quite large or impinge on nerves of the
pelvic limbs. Smaller tumors may be detected by coelomic palpation. On
exploratory celiotomy or postmortem examination, tumors may be single
or multiple and typically are white, slightly rm, and smooth or
multinodular (Fig. 5A). Larger tumors often will have central necrosis
and regional visceral adhesions. Histologically, Lucke tumors typically are
well demarcated, and comprised of neoplastic tubules that may be ectatic
or lined by papillary projections. Intranuclear inclusion bodies are
numerous in the algid phase and rare in the calid phase [43,49]. Also,
calid phase tumors can have a high mitotic rate, consistent with rapid
growth [14]. Tumors may arise anywhere within the kidneys and aect
both kidneys similarly. A slight predisposition for the right kidney is
reported [50]. Metastasis can occur to any organ, especially the liver [14].
The reproductive tract may be intimately associated with or compressed by
the tumor, requiring careful dissection.
Diagnosis is based on the species and presence of renal tumor(s).
Dierential diagnoses include other causes of coelomic distension, such as
eusion, coelomitis, parasitism, and egg production. Renal granulomas,
such as from mycobacteriosis, should be considered with smaller masses
(Fig. 5B). A ne-needle aspirate or touch preparations may facilitate antemortem diagnosis. Neoplastic epithelial cells exfoliate well and often are
easily identiable.
Other renal tumors
Other types of renal tumors are rare and include single case reports of
nephroblastoma and renal carcinoma in various species of anurans and
urodeles [14,51,52]. One exception is a high prevalence of polycystic kidneys
in a population of hybrid Japanese toads (Bufo japonicus) and Chinese toads
(Bufo raddei). The disease appears to follow a progression from hyperplasia
of the tubular epithelium to tubular or papillary renal adenocarcinomas
[53]. These tumors are believed to have a genetic basis, and in some cases,
metastasized to the lungs, liver, and kidneys.
Testicular tumors
Amphibian testes are internal and undergo seasonal variation in
spermatogenetic activity. Reports of neoplasia are limited to Sertoli cell
tumors in both anurans and urodeles. These tumors are considered an
incidental nding at necropsy. A series of testicular tumors reported in 16 A

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Fig. 5. (A) A Northern leopard frog (Rana pipiens) with a large Lucke adenocarcinoma of the
right kidney that presented with marked coelomic distention. This is a large calid phase tumor
and metastases to the liver and lungs was noted on histologic examination. Lucke tumors were
subsequently diagnosed in ve out of 10 animals in this group. (B) Coelomic viscera from a R
pipiens (left) and Tropical clawed frog (Xenopus (Silurana) tropicalis) (right) demonstrating the
similar gross appearance of neoplasia and granulomatous inammation. The R pipiens kidneys
are expanded by multiple Luckes adenocarcinomas. One cranial pole of the X tropicalis kidney
is eaced by granulomatous inammation due to mycobacteriosis.

mexicanum by Humphrey [54] were classied as Sertoli cell tumors in the

RTLA [14,54]. All of these cases occurred in a single colony, and 14 of the
animals were closely related, which suggests a hereditary predisposition in
this group. Many of these tumors were pedunculated and white with deep
ssures and a pebbled appearance. Neoplastic cells formed elongated,
branched tubules. Atrophy of the preexisting testicular tissue was noted in
some cases. All of the tumors were benign and nonfatal. Sertoli cell tumors
also are described in a Hellbender (Cryptobranchus alleganiensis) and a R
pipiens  R palustris hybrid [14,55]. Seminomas and interstitial (Leydig) cell
tumors are not reported.

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Ovarian tumors
Ovarian tumors are poorly described in amphibians. Many are identied
simply as epithelial neoplasia or carcinomas. Use of nonspecic classication schemes makes it dicult to draw rm conclusions regarding specic
tissue of origin, biologic behavior, and similarities with primary ovarian
tumors in other species. In other species, the term epithelial usually
implies an origin from the surface epithelium of the ovary. Canine ovarian
cystadenomas and cystadenocarcinomas are relatively common examples.
One cystadenocarcinoma was reported in a R pipiens [56]. Green and
Harshbarger [14] report a single case of a granulosa cell tumor in an ornate
horn frog, and Streett [57] reports an ovarian teratoma in R pipiens.
Dysgerminomas are not documented in amphibians.

Endocrine organs
Neoplasia of endocrine organs is reported rarely. With the exception of
the Asian frog population described below, endocrine tumors are limited to
a presumed adenoma of the interrenal gland in R pipiens and suspect
pancreatic carcinoma in X laevis [14].
Pancreatic carcinoma of Asian pond frogs
Masahito et al [40] described over 101 tumors mostly of pancreatic origin
in a captive population that included multiple species of pond frogs
originating from Japan (Rana nigromaculata and R porosa brevipoda),
China (R nigromaculata and R plancyi plancyi), Korea (R nigromaculata),
and Taiwan (R plancyi fukienensis). Tumors ranged from 0.5 to 3.7 cm in
diameter, expanding the region of the pancreas. Most tumors originated
from the endocrine (islet) cells with typical histomorphology of tightly
packed groups of nests, rudimentary tubules, and acini. Areas of exocrine
dierentiation were noted in some tumors, which also is a feature of some
islet tumors in mammalian species [58]. Immunohistochemical characterization of a subset of cases revealed expression of insulin or somatostatin in
many tumors. Metastases were observed in the liver, kidney, heart, lung,
stomach, and ovary.
Tumors were observed rst in hybrids bred from the imported Chinese
frogs, and eventually in other hybrid and nonhybrid frogs. Subsequent
examination by transmission electron microscopy revealed type C retrovirus
particles in the tumor cells. The epidemiology and presence of viral particles
in these tumors suggest a viral etiology. Two other types of tumors were
identied in this group including a rhabdomyosarcoma and a hepatic
adenoma. It is not known whether the etiology of these tumors was
associated with the pancreatic carcinomas.

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Pseudoneoplastic conditions
In general, the majority of masses of amphibians are nonneoplastic and
caused by bacteria, fungi, parasites, or other infectious or inammatory
conditions. Here we review two commonly encountered pseudoneoplastic
conditions, mycobacteriosis, and oral inammatory lesions.
Mycobacteriosis
Mycobacteriosis is a common disease of captive anurans, and can cause
a variety of clinical signs. Multiple mycobacterial species are demonstrated
as causative agents. Currently, it is considered an incurable disease in
amphibians and its eradication from a collection is problematic.
Granulomatous inammation caused by mycobacteria has been misidentied on gross and histologic evaluation as various types of neoplasia,
including lymphoma and mesenchymal tumors, and often requires histopathology for denitive diagnosis (Fig. 6). Misdiagnosis of mycobacterial
lesions is the source of great confusion in the literature. Reports and
experimental studies of lymphoma in X laevis and T pyrrhogaster ultimately
were shown to be mycobacteriosis [59,60]. During the investigation of this
disease, various hypotheses were proposed, such as secondary infection of
tumors by mycobacteria and mycobacteria-induced neoplastic transformation [59,61]. Eventually, most investigators disregarded these hypotheses
[60,62]. Certain features of mycobacterial lesions contribute to misinterpretation, especially by individuals not familiar with amphibian pathophysiology and histology. Namely, macrophages and reactive broblasts of
granulomas were misidentied as sarcomas or lymphomas. Also, myeloid
hyperplasia of the hepatosplenic hematopoietic tissue is a cellular response
in infectious processes, such as mycobacterial infections, and can be
misinterpreted as lymphoma. Histologically, there are various degrees of
proliferation of hematopoietic tissue in the periportal and subcapsular
regions of the liver and spleen. The reactive cell population is pleocellular,
with a progression of leukocyte development that may include a large
number of blastic precursor cells (Fig. 7). This response is described in the
literature and is observed commonly in Xenopus species and A mexicanum at
UCB that are infected with mycobacteria and, less frequently, other bacteria
species such as Chryseobacterium (Flavobacterium). Myeloid hyperplasia
may occur in the absence of hepatic granulomas or other visceral
involvement, such as in some cases of cutaneous mycobacteriosis. Misdiagnosis can be avoided by careful histologic examination and prudent use
of acid-fast stains.
Inammatory lesions and myxoid tumors
Veterinarians have encountered proliferative, mass-like lesions protruding from the oral cavity, characterized by a prominent myxoid or mucinous

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Fig. 6. Ovaries, oviducts, and kidneys (center) from an African clawed frog (Xenopus laevis)
demonstrating the pseudoneoplastic appearance of granulomatous inammation. The
abnormal smooth tissue (arrows) expands and obliterates the normal ovary (asterisks), as well
as the kidneys, and corresponds to areas of intense granulomatous inammation. This is
another example of mycobacteriosis and diagnosis required histological examination.

matrix. Anecdotal evidence indicates that these lesions occur with some
frequency, especially in the maxilla or other regions of the oral cavity. A few
such cases have been reported in the literature and accessioned into the
RTLA [14]. Diagnoses such as myxoma, myxosarcoma, and chondromyxoma are applied; however, neoplastic origin is not proven. Green and
Harshbarger [14] comment on the uncertain etiology and consider dierential causes, such as chronic, exuberant inammation, and callus formation.
One case encountered at UCB responded well to surgical excision, and
histologic examination revealed features consistent with a chronic, reactive
lesion, including inammation, broplasia, and periosteal response (Figs.
8A and 8B). More work is needed to characterize these conditions.

Treatment
Treatment options for neoplasia are limited to removal of predisposing
condition(s), supportive care, and surgical excision. To the best of the
authors knowledge, no studies have examined the ecacy of chemotherapy
in amphibians. As with many physiologic processes in ectotherms, the
growth of some types of neoplasia is temperature dependent. Thus, ambient
temperature can inuence growth rate, treatment, and outcome, and may be
manipulated to inuence biologic behavior and tumor incidence. When
treatment options fail or are not suitable, humane euthanasia may be
elected.
Many tumors of the integument in amphibians are limited to the dermis,
and those aecting anurans are often amenable to surgical excision.

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Fig. 7. Photomicrograph of the liver of a Tropical clawed frog (Xenopus tropicalis) with
mycobacteriosis. Hepatic cords (asterisks) are separated by proliferating myeloid cells (M)
(myeloid hyperplasia), including mitotically active progenitor cells (arrow). This reactive change
is commonly encountered in amphibians with mycobacteriosis and may be misinterpreted as
lymphoproliferative disease. Hematoxylin and eosin; 400.

Furthermore, amphibians are excellent surgical candidates due to their

remarkable capacity for regeneration, which varies among species. Urodeles, newts in particular, have remarkable regenerative abilities of the
appendages and tail [63]. In the authors experience, there are marked
dierences in the degree of technical diculty between surgical procedures
performed on anurans and urodeles. Anurans have highly vascularized,
pliable, loosely attached integument that is quite amenable to surgical
manipulation. Conversely, the skin of urodeles is adhered rmly to the deep
muscular layer, making wound closure more dicult. A complete review of
amphibian anatomy is beyond the scope of this article; however, before
performing any major surgery, a review of vital structures, such as lymph
hearts and the midline coelomic vein, is recommended.
Basic surgical procedure
In amphibians, the most common anesthetic protocol for induction is
immersion in a known concentration of tricaine (MS222). Benzocaine, clove
oil, and volatile anesthetics may be suitable alternatives, and are imprecisely
administered to eect. Gauze soaked in sterile water should be placed
underneath the patient to maintain cutaneous moisture. Information on
postoperative pain management is limited. A few studies suggest that
opiods, a-adrenergic, and nonsteroidal antiinammatory agents may provide pain relief [64,65].
Surgical procedures on amphibians are considered clean contaminated.
Sterile preparation of the surgical site is accomplished by placing gauze
soaked in dilute chlorhexidine solution (0.75%) or benzalkonium solution (2
mg/L) on the surgical site for 10 minutes. Other disinfectants commonly

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Fig. 8. (A) Two-year-old Pacic tree frog (Hyla (Pseudacris) regilla) with a large, multinodular,
eshy mass originating from the ventral aspect of the rostral maxilla. The mass was successfully
excised, and the defect was allowed to heal by contraction and epithelialization. (B)
Photomicrograph of the maxillary mass from the Hyla regilla in (A). The mass is comprised
of reactive broblasts and abundant mucinous stroma (asterisks). Inammatory cells (arrows)
diusely inltrate the tissue. These histologic features are consistent with a reactive
inammatory process. Hematoxylin and eosin; 20 (inset 400).

used for mammals, such as iodine compounds or alcohol-based solutions,

should be avoided due to possible toxic aects. In anurans but not urodeles,
the skin is tented easily to make a stab incision. Once a window is created,
the deep surface of the skin can be inspected visually to avoid transection of
major vessels. An excisional tumor margin of at least 0.2 to 1.0 cm is
recommended, but will be limited by species, size, and location. Skin closure
is performed using nonabsorbable, monolament suture in an interrupted
everting pattern. In several anuran species, extensive wound management is
not required because of inherent antimicrobial peptides that are excreted
from the skin [6668]. Hygienic and optimal environmental conditions,
however, are required in the postoperative interval to avoid opportunistic
bacterial and fungal infections.

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Summary
This article is a review of the more common types of neoplasia in
amphibians. When possible, clinically relevant features, such as biologic
behavior, anatomy, associated etiologies, and major dierential diagnoses,
are included. It is important to recognize neoplasia caused by oncogenic
viruses or predisposing conditions, and to distinguish neoplasia from
infectious pseudoneoplastic conditions that can impact population and
group health. Treatment options are limited to removal of identied
predisposing condition(s), supportive care, controlling ambient temperatures, surgical excision and, when necessary, humane euthanasia.

Acknowledgments
We would like to thank Drs. Hilde E. V. De Cock, Helen E. Diggs, Nina
Hahn, John Harshbarger, Tyrone Hayes, Linda J. Lowenstine, Yuka
Nakamura, Allen Pessier, Francis Rose, Gregory Timmel, and Tanja S.
Zabka for their editorial support, consultation, and contribution of cases.
We also thank Jerry Fields (UCD) for his assistance with photography;
Stephen Friet, Sam Cunningham, Loren Johnson, Jeanine Hamel, Rachel
Casamina, Aaron Vonk, and Magdalena Mleczko of the UC Berkeley,
Oce of Laboratory Animal Care; and Barry Puget, Asi Djan, and Diane
Naydan of the UC Davis histology laboratory.
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