VDE_301.

fm Page 243 Friday, September 20, 2002 1:09 PM

Veterinary Dermatology 2002, 13, 243 246

Case report

Blackwell Science, Ltd

Primary cutaneous extragenital canine transmissible venereal

tumour with Leishmania-laden neoplastic cells: a further suggestion
of histiocytic origin?
Histiocytic origin of TVT

FRANCESCO ALBANESE,* ALESSANDRO POLI, FRANCESCA MILLANTA and

FRANCESCA ABRAMO
*Practitioner, Naples, Italy;
Department of Animal Pathology, University of Pisa, Pisa, Italy
(Received 11 June 2001; accepted 22 April 2002)

Abstract The clinical signs and histopathological features of a primary extragenital canine transmissible
venereal tumour (TVT) are described. Three subcutaneous round alopecic nodules were located on the anterior
and caudal dorsal region and in the ventral area of the neck. Cytologically, tumour cells were intermediate in
size with a moderate amount of cytoplasm, and the nuclei were immature with finely reticular chromatin. The
cytoplasm was lightly to heavily basophilic and contained distinct small vacuoles at the periphery. On the basis
of these characteristics, a diagnosis of TVT was made and confirmed by histological and ultrastructural investigations. Leishmania amastigotes were detected in the cytoplasm of macrophages and neoplastic cells of the
tumoral mass. The presence of the parasite within neoplastic cells is consistent with a histiocytic origin of TVT.
Keywords: Leishmania amastigotes, nodule, skin, transmissible venereal tumour.

INTRODUCTION
Transmissible venereal tumour (TVT) is a transplantable
tumour unique to the dog. Owing to the nature of
transmission by sexual contact, the external genitalia of
either sex are the primary site of tumour involvement.
Particular social behaviour, such as sniffing, licking
or scratching, can lead to the presence of extragenital
implants that generally occur in the nasal or oral cavity.1
In the literature, the simultaneous occurrence of the
tumour in the genitalia, the skin, the subcutaneous
tissue, the internal organs and the central nervous
system has been reported.24 Primary cutaneous extragenital localization of the tumour is seldom documented.57 Under experimental conditions, the clinical
course of the illness is affected by the animals immune
response.8 This is supported by the fact that stray dogs
with a low standard of care or immunosuppressed
dogs are more susceptible. Immunocompromised dogs
are also more prone to metastatic lesions.911
Leishmaniasis caused by the intracellular protozoan
Leishmania infantum is an endemic disease in the
Mediterranean basin. The presence of parasitized
macrophages in affected dogs has suggested impaired
cellular immunity which can favour the dissemination
of the parasite.1215
Correspondence: Francesca Abramo, Department of Animal Pathology, Viale delle Piagge, 2, I-56124, Pisa, Italy. Fax: +39-50-560544;
E-mail: abramo@vet.unipi.it
2002 Blackwell Science Ltd

This study describes a case of a multiple primary

extragenital cutaneous TVT in a dog, in association
with L. infantum infection. The presence of the parasite
both in the macrophages and the TVT neoplastic cells
suggests the histiocytic origin of the neoplasm.

CASE REPORT
A 7-year-old, intact female, mixed-breed dog from a
kennel was presented for the presence of cutaneous
nodules. Upon physical examination, the dog was
found to be severely depressed, anorexic and in a poor
general health. External mucosae were pale and all
peripheral lymph nodes were enlarged slightly. Dermatological examination showed three subcutaneous
rounded alopecic masses 68 cm in diameter, located
in the anterior and caudal dorsal region, as well as the
ventral area of the neck. The nodules had appeared in
the last 3 weeks. A seborrheic dermatosis was observed
over the entire body, especially affecting the dorsum,
and over the nodules. Additional investigations were
undertaken and routine haematological analysis and
blood chemistry demonstrated (normal ranges given
in parentheses): red cell count, 3.11 1012/L (5.47.8);
haematocrit, 19% (3754); haemoglobin, 67 g/ L (128
180); white cell count, 8.8 109/L (6.017.0); platelet
count, 392 109/L (120350); urea nitrogen, 22.1
mmol/L (1.54.5); creatinine, 460 mol/L (30170);
total protein, 98 g/L (5577); albumin, 18 g/ L (2540);
243

VDE_301.fm Page 244 Friday, September 20, 2002 1:09 PM

244

F. Albanese et al.

Figure 1. Canine transmissible venereal tumour. Individually

exfoliating round cells characterized by nucleus with prominent
nucleolus and presence of variable number of small vacuoles in the
cytoplasm adjacent to the distinct cell boundary (Diff
Quick, 1300).

Figure 2. Canine transmissible venereal tumour. Round cells filled

with Leishmania amastigotes; notice numerous intracytoplasmic
amastigotes with flagellar remnants (Diff Quick, 1000).

globulin, 80 g/L (2545); alanine aminotransferase,

98 U/L (065); and aspartate aminotransferase 88 U/L
(067). Urine analysis showed a moderate proteinuria
(> 1000 mg/L). Ecographic evaluation revealed a mild
splenomegaly and the absence of internal masses. The
presence of serum anti-Leishmania antibodies was determined by the indirect immunofluorescence test (IFT) as
described previously,16 and a titre of 320 was recorded.
Fine-needle aspirates taken from the cutaneous
nodules were air-dried, stained with Diff-Quick and
examined cytologically. The monomorphous cell
populations consisted mainly of discrete round cells
with clear or slightly basophilic cytoplasm containing
numerous clear small vacuoles arranged in chains or a
horseshoe pattern around the cell boundary (Fig. 1).
Nuclei contained clumped chromatin, one or more
prominent nucleoli and abnormal mitoses were present.
Macrophages, lymphocytes and plasma cells were also
detected. Intracytoplasmic L. infantum amastigotes
were observed in the infiltrating macrophages and in
the neoplastic cells (Fig. 2). Cytological findings were
consistent with the diagnosis of concurrent TVT and
leishmaniasis.
2002 Blackwell Science Ltd, Veterinary Dermatology, 13, 243 246

Figure 3. Canine transmissible venereal tumour. Solid sheets of

round cells with clear cytoplasm, separated by collagen bundles.
Several mitoses are present (H&E stain, bar = 25 m).

Excisional biopsies from the three cutaneous nodules

were collected. Representative portions were fixed in
10% neutral buffered formalin solution (pH 7.4) and
embedded in paraffin wax. Four-micrometre sections
were stained with haematoxylin and eosin (H&E) and
Giemsa stains. In all sections the neoplastic cells were
arranged in solid sheets, clusters or cords interlaced
by a delicate vascular stroma. The round neoplastic cells
had abundant cytoplasm that was either clear or finely
granular. Nuclei showed chromatin clumping and
contained one or more prominent nucleoli (Fig. 3);
in most cases atypical mitotic figures were present.
Mitoses ranged from 1 to 4 per microscopic field at 400
magnification. The cells had a large nuclear/cytoplasmic
ratio. Focally, a variable number of intracytoplasmic
amastigotes was present in vacuolated neoplastic cells
ranging in number from 1 to 3 per cell.
Representative 1 mm3 pieces from one of the formalin
fixed nodule were washed in phosphate-buffered saline
(PBS, pH 7.4) and processed for transmission electron
microscopy. Briefly, the pieces were fixed in a 2.5%
glutaraldehyde solution in 0.1 PBS, postfixed in 1%
buffered osmium tetroxide and embedded in epoxy
resin. Ultrathin sections were double stained with 2%
uranyl acetate and lead citrate and examined using
a Philips CM-10 transmission electron microscope. A
monomorphous population of closely packed round
cells and infiltrating inflammatory cells was present. The
round cells had abundant cytoplasm containing rough
endoplasmic reticulum and mitochondria of varying
size and shape. A variable number of electron-lucent
vacuoles was found adjacent to the cell membrane.
Tight junctions were not observed and cell boundaries
were distinct with extensive interdigitation between
the cell membranes of adjacent cells (Fig. 4). Round
cells also contained a variable number of 13 m long
intracytoplasmic amastigotes (close to the vacuoles)
characterized by a central nucleous and a kinetoplast.
Infiltrating inflammatory cells were mainly macrophages,
most of which contained intracytoplasmic parasites,
lymphocytes and plasma cells.

VDE_301.fm Page 245 Friday, September 20, 2002 1:09 PM

Histiocytic origin of TVT

Figure 4. Canine transmissible venereal tumour cell containing a

large nucleus with prominent nucleolus. The cytoplasm contains
several vacuoles and a Leishmania amastigote. Cell boundaries are
characterized by distinct interdigitation without intercellular bridges
(uranyl acetate-lead citrate stain, 4400).

245

rectal involvement together with a vaginal TVT was

present in another case. Consequently, cutaneous primary extragenital sites seem to be very rare.57 Most
of these cases were diagnosed on formalin-fixed tissue
and not by impression smears. In this case study, the
diagnosis was made by cytology and the coexistence
of Leishmania amastigotes in TVT nodules has been
reported for the first time. Gross and histological
lesions observed in kidneys and gross alterations of the
spleen were compatible with visceral leishmaniasis.18
The expression of class II MHC antigens by canine
TVT cells8,19 has suggested a reticuloendothelial origin
of the tumour. Neoplastic cells also express lysozyme
and alfa-1-antitrypsin immunoreactivity20 and a canine
macrophage marker.21 In our case, Leishmania amastigotes were located in the cytoplasm of both infiltrating
macrophages and neoplastic cells, further supporting the
monocyte/macrophage lineage histogenesis of the TVT.

REFERENCES
In order to avoid renal complications, the subject
was given fluid therapy. Nasogastric tube feeding,
an hydrating solution and erythropoietin 100 UI/kg
subcutaneously every two days, were also administered.
The subject was also given an intravenous injection of
0.075 mg/kg vincristine and after one week the nodules
decreased in size. However, because of a progressive
worsening of the general condition of the dog, euthanasia
became necessary. At necropsy, neither genital nor
internal masses were revealed, the spleen was enlarged
and the kidneys were slightly pale. Histological examination of kidneys showed evidence of membranoproliferative glomerulonephritis, interstitial periglomerular
and peritubular lymphoplasmacytic nephritis.

DISCUSSION
As the genital lesions of TVT are due to the coitus,
extragenital cases of TVT occurring in the nasal and
oral cavity are usually associated with particular social
behaviour. In the case of simultaneous localization of
the tumour in the genitalia and the skin, it is difficult
to distinguish between a primary cutaneous extragenital
location and a metastatic site, as metastasis of the TVT
can occur even if rarely (017%).4,7,17 In this case a
primary extragenital cutaneous TVT is documented.
No primary tumour was detected in the genital area
or in the oral or nasal cavity, making metastatic dissemination unlikely. Moreover, males are considered to be
more susceptible to metastatic diffusion than females,
as reported by some authors.11 In a recent retrospective
study, 93.7% of 29 TVT were located in the genital area
and the remainder in the nasal cavities. One of these
dogs also had cutaneous nodules in the ventral cervical
region.3 Other authors reported 18 cases of TVT of
which solitary primary genital lesions were diagnosed
in 16 dogs. Three cutaneous nodules were present
in addition to a tumour on the penis in one case, and

1. Weir, E.C., Pond, M.J., Duncan, J.R. et al. Extragenital

occurrence of TVT in the dog: literature review and
case reports. Journal of the American Animal Hospital
Association 1978; 14: 5326.
2. Oduye, O.O., Ihede, B.O., Esuruose, G.E. et al. Metastatic TVT in dogs. Journal of Small Animal Practice
1973; 14: 62537.
3. Rogers, K.S., Walker, M.A., Dillon, H.B. Transmissible
venereal tumour: a retrospective study of 29 cases.
Journal of the American Animal Hospital Association
1998; 34: 46370.
4. Ferreira, A.J., Jaggy, A., Varejao, A.P. et al. Brain and
ocular metastases from a transmissible venereal tumour
in a dog. Journal of Small Animal Practice 2000; 41:
1658.
5. Feldman, W.H. So-called infectious sarcoma of the dog
in an unusual anatomic situation. American Journal of
Pathology 1929; 5: 18395.
6. Abbott, P.K. Venereal transmissible tumour on eyelid of
dog. Australian Veterinary Journal 1966; 42: 29.
7. Higgins, D.A. Observations on the canine transmissible
venereal tumour as seen in the Bahamas. Veterinary
Record 1966; 79: 6771.
8. Cohen, D. The canine transmissible venereal tumour: a
unique result of tumour progression. Advances in Cancer
Research 1985; 43: 75112.
9. Cohen, D. The biological behaviour of the transmissible
venereal tumour in immunosuppressed dogs. European
Journal of Cancer 1973; 9: 2538.
10. Amber, E.I., Henederson, R.A., Adeyanju, J.B. et al.
Single-drug chemotherapy of canine transmissible
venereal tumour with cyclophosphamide, inethorrexate,
or vincristine. Journal of Veterinary Internal Medicine
1990; 4: 1407.
11. Boscos, C.M., Tontis, D.K., Samartzi, F.C. Cutaneous
involvement of TVT in dogs: a report of two cases.
Canine Practice 1999; 24: 611.
12. Carvalho, E.M., Teixeira, R.S., Johnson, W.D. Cellmediated immunity in American visceral leishmaniosis:
reversible immunosuppression during acute infection.
Infectious Immunology 1981; 33: 498502.
2002 Blackwell Science Ltd, Veterinary Dermatology, 13, 243246

VDE_301.fm Page 246 Friday, September 20, 2002 1:09 PM

246

F. Albanese et al.

13. Keenan, C.M., Hendricks, L.D., Lightner, L. et al.

Visceral leishmaniosis in the German Shepherd dog. I.
Infection, clinical disease and clinical pathology. Veterinary Pathology 1984; 21: 74 9.
14. Rhalem, A., Sahibi, H., Guessous-Idrissi, N. et al.
Immune response against Leishmania antigens in dogs
naturally and experimentally infected with Leishmania
infantum. Veterinary Parasitology 1999; 81: 17384.
15. Leandro, C., Santos-Gomes, G.M., Campino, L. et al.
Cell mediated immunity and specific IgG1 and IgG2
antibody response in natural and experimental canine
leishmaniasis. Veterinary Immunology and Immunopathology 2001; 79: 273 84.
16. Mancianti, F., Meciani, N. Specific serodiagnosis of canine
leishmaniasis by indirect immunofluorescence, indirect
hemagglutination and counter-immunoelectrophoresis.
American Journal of Veterinary Research 1988; 49:
1409 11.

17. Rogers, K.S. Transmissible venereal tumour. Continuing

Education Article 1997; 19: 103645.
18. Poli, A., Abramo, F., Mancianti, F. et al. Renal
involvement in canine leishmaniasis. A light microscopic,
immunohistochemical and electron-microscopic study.
Nephron 1991; 57: 44452.
19. Epstein, R.B., Bennet, B.T. Histocompatibility typing
and course of canine venereal tumours transplanted into
unmodified random dogs. Cancer Research 1974; 34:
78893.
20. Mozos, E., Mendez, A., Gomez-Villamandos, J.C. et al.
Immunohistochemical characterisation of canine
transmissible venereal tumour. Veterinary Pathology
1996; 33: 25763.
21. Marchal, T., Chabanne, L., Kaplanski, C. et al. Immunophenotype of the canine transmissible venereal tumour.
Veterinary Immunology and Immunopathology 1997; 57:
111

Rsum Cet article dcrit les signes cliniques et histopathologiques dune localisation extragnitale dune
tumeur vnrienne transmissible (TVT) . Lanimal prsentait trois nodules sous-cutans de forme ronde, localiss
sur un antrieur, la rgion caudale dorsale, et la face ventrale du cou. Lexamen cytologique montrait des cellules
tumorales de taille intermdiaire, avec un cytoplasme peu abondant et un noyau immature chromatine finement
rticule. Le cytoplasme tait modrment fortement basophile, et contenait des petites vacuoles en priphrie.
Sur la base de ces lements, un diagnostic de TVT a t fait, ce qui a t confirm par les examens histologiques
et ultrastructuraux. Des amastigotes de Leishmania ont t dtects dans le cytoplasme des macrophages et des
cellules noplasiques dans la tumeur. La presence de parasites lintrieur des cellules noplasiques est en faveur
dune origine histiocytaire de la TVT.
Resumen Se describen los sntomas clnicos y las caractersticas histopatolgicas de un tumor venreo transmisible (TVT) primario extragenital. Tres ndulos subcutneos, redondos y alopcicos se encontraban localizados
en la regin dorsal anterior y caudal, y en el rea ventral del cuello. Citolgicamente las clulas tumorales eran
de tamao intermedio, con una cantidad moderada de citoplasma, y los ncleos eran inmaduros con cromatina
discretamente reticular. El citoplasma mostraba una basoflia leve a intensa y contena pequeas vacuolas
evidentes en la periferia. Basndonos en estas caractersticas se estableci un diagnstico de TVT y se confirm
mediante estudios histolgicos y ultraestructurales. Se detectaron amastigotes de Leishmania en el citoplasma
de macrfagos y clulas neoplsicas de la masa tumoral. La presencia del parsito en clulas neoplsicas es
compatible con un origen histioctico del TVT.
Zusammenfassung Es werden die klinischen und histopathologischen Eigenschaften eines primren, extragenitalen caninen Transmissiblen Venerischen Tumors (TVT) beschrieben. Drei subkutane, runde und alopezische
Knoten befanden sich in der cranialen und caudalen Rckenregion sowie in der ventralen Region des Halses.
Zytologisch waren die Tumorzellen von mittlerer Gre mit mittelgroem Zytoplasma und besaen einen
unreifen Nukleus mit feinem retikulrem Chromatin. Das Zytoplasma war gering- bis hochgradig basophil und
enthielt in der Peripherie kleine Vakuolen. Anhand dieser Charakteristika wurde die Diagnose eines TVT gestellt
und anschlieend mit Hilfe histologischer und ultrastruktureller Untersuchungen besttigt. Leishmania Amasigoten wurden im Zytoplasma von Makrophagen sowie neoplastischer Zellen entdeckt. Die Anwesenheit dieses
Parasits innerhalb der neoplastischen Zellen ist vereinbar mit der histiozytren Herkunft des TVT.

2002 Blackwell Science Ltd, Veterinary Dermatology, 13, 243 246