Cytological and Histopathological Evaluation of Canine Transmissible Venereal Tumour (CTVT)

Introduction

Transmissible venereal tumour (TVT) is one of the most uncontrolled tumours in dogs (Ucar, 2016) [15] . It is one of the most common benign tumours in dogs that can be seen in both genders (Kucukberkir et al., 2021) [6] . Canine Transmissible Venereal Tumour (CTVT) is a naturally occurring, clonally transmissible round cell neoplasm of dogs, transmitted primarily through direct contact during coitus and occasionally by social behaviours such as licking or sniffing of affected areas. It is one of the few known naturally transmissible cancers and is widely prevalent in tropical and subtropical regions, particularly among free-roaming and sexually active dogs (Nowinsky, 1876; Murgia et al., 2006) [12,11] . Clinically, CTVT commonly affects the external genitalia of both male and female dogs and presents as a friable, haemorrhagic, cauliflower-like mass associated with vaginal or preputial bleeding (Das and Das, 2000; Purohit, 2009) [1,13] . Although gross appearance often suggests the diagnosis, confirmation is essential to differentiate CTVT from other genital neoplasms and inflammatory conditions. CTVT, macroscopically, can be solitary or multiple tumour masses with cauliflower-like ulceration, haemorrhagic, friable and irregular appearance can be seen. Lesions are fragile and mostly haemorrhagic as a result of low cohesion amongst neoplastic cells. Tumour size varies from millimeters (mm) to several centimeters (less than 1 to 3 mm in diameter) with a dark red to greyish pink colour (Das and Das, 2000; Purohit, 2009; Lopes et al., 2015) [1,13,15] . As microscopically diagnosis is made by histopathological examination of the biopsy specimen, large cells with round or oval vague contours are observed on histological examination. CTVT is morphologically categorised as a spherical (round) cell tumour, however, although the precise origin of the cells is unknown, immunohistochemical investigations revealed histiocytic and mesenchymal origin (Do Amaral et al., 2007) [2] . Cytologically, CTVT cells are round or oval, 14 to 30 μm in diameter, with pale blue cytoplasm which is particularly vacuolated, with a relatively elevated nuclear: cytoplasmic ratio. The cells also comprise dense chromatin, visible nucleoli and numerous mitotic structures, as well as anisocytosis and anisokariosys (Erünal-Maral et al., 2000) [3] .

Cytological techniques such as fine needle aspiration cytology (FNAC) and impression smear are widely used as rapid, minimally invasive, and cost-effective diagnostic tools for CTVT. These techniques allow early identification of characteristic tumour cells with high diagnostic accuracy (Erünal-Maral et al., 2000; Ganguly et al., 2016) [3,4] . However, histopathological examination remains the gold standard for definitive diagnosis, as it provides detailed information regarding tissue architecture, cellular arrangement, mitotic activity, and stromal response (Mukaratirwa and Gruys, 2003) [9] . Therefore, combined cytological and histopathological evaluation is recommended for accurate diagnosis of CTVT.

Materials and Methods

Dogs presented with clinical signs suggestive of CTVT, such as genital bleeding and visible tumour masses, were included in the study. All cases were subjected to confirmatory diagnostic evaluation prior to initiation of therapy. Dogs under Group 1 (6 animals) were treated with vincristine sulphate at the dose rate of 0.025 mg/kg body weight intravenously, once a week, for 3 to 5 weeks depending upon the regression of tumour mass. Dogs under Group 2 (6 animals) were treated with bacillus Calmette-Guerin (BCG) at the dose of 2-8 × 10⁶ CFU dissolved in 5.0 ml of normal saline. The solution was administered intratumorally and also applied as a local spray once a week for 3 to 5 weeks. Dogs under Group 3 (6 animals) were treated with combination therapy of vincristine sulphate and BCG using the same dose and protocol as described for Groups 1 and 2.

Diagnosis of TVT was confirmed by impression smear, fine needle aspiration cytology (FNAC) and histopathological examination. FNAC was performed by aspirating tumour tissue using a fine gauge needle under aseptic conditions. The aspirated material was smeared onto clean glass slides, airdried, and stained using routine cytological stains. The smears were examined under light microscopy for cellular morphology and arrangement. Impression Smear Impression smears were prepared by gently pressing the freshly cut surface of the tumour mass onto clean glass slides. The smears were air-dried, stained, and examined microscopically to evaluate cytological features. Histopathological Examination Tumour biopsy samples were collected from representative areas of the mass. The tissues were fixed in 10% neutral buffered formalin, processed by routine paraffin embedding, sectioned at 4-5 μm thickness, and stained with Haematoxylin and Eosin (H&E). Histological evaluation focused on tumour cell morphology, mitotic activity, stromal response, and inflammatory cell infiltration.

Results

FNAC smears revealed high cellularity with numerous round to polyhedral neoplastic cells arranged in sheets and rows. The tumour cells showed moderate to abundant pale basophilic cytoplasm with distinct cytoplasmic vacuolation. Nuclei were round to oval, hyperchromatic, and exhibited prominent nucleoli with a high nuclear-to-cytoplasmic ratio. Frequent mitotic figures and cellular pleomorphism were observed. Inflammatory cells, mainly lymphocytes and neutrophils, were also present in the background.

Discussion

The present study demonstrates that FNAC, impression smear, and histopathological examination are valuable diagnostic modalities for the diagnosis of canine transmissible venereal tumour. FNAC and impression smear provided rapid and reliable preliminary diagnosis by revealing characteristic round to polyhedral tumour cells with vacuolated cytoplasm, high nuclear-to-cytoplasmic ratio, hyperchromatic nuclei, and prominent nucleoli. Similar cytological features have been reported as diagnostic hallmarks of CTVT by Erünal-Maral et al. (2000) [3] and Ganguly et al. (2016) [4] . FNAC yielded better cellularity and clearer nuclear details compared to impression smear, making it a preferred cytological technique, which is in agreement with earlier reports (Gurel et al., 2002; Lather et al., 2017) [5,7] . Impression smear, although simple and quick, lacked consistency in cellular yield and did not provide information on tissue architecture. Histopathological examination confirmed the diagnosis by demonstrating sheets and cords of round to polyhedral tumour cells supported by delicate connective tissue stroma, along with high mitotic activity and inflammatory cell infiltration. These findings are consistent with the classical histological description of CTVT reported by Mukaratirwa and Gruys (2003) [9] and Stockmann et al. (2011) [14] . Histopathology also allows differentiation of CTVT from other round cell tumours and provides essential baseline data for assessment of tumour behaviour and therapeutic response. Thus, while cytological techniques are useful for rapid screening and field diagnosis, histopathology remains indispensable for confirmatory diagnosis. A combined diagnostic approach enhances accuracy and reliability, as supported by previous studies (Ganguly et al., 2016; Mukaratirwa et al., 2009) [4] .

Conclusion

FNAC and impression smear are effective, rapid, and minimally invasive diagnostic techniques for the preliminary diagnosis of canine transmissible venereal tumour. Histopathological examination remains the gold standard for definitive diagnosis due to its ability to assess tissue architecture and cellular characteristics. A combined diagnostic approach using cytology and histopathology enhances diagnostic accuracy and supports effective clinical management of CTVT in dogs.

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