An investigation of bovine papillomaviruses from ocular squamous cell carcinomas in cattle

Abstract

Background: Ocular squamous cell carcinomas (OSCCs) in cattle has been studied for many years, but no definite etiology has been established. Squamous cell carcinomas (SCCs) may occur in different body parts of cattle. Depending on the location, it can cause an economic loss of varying degrees.

Aims: The aim of this study was to investigate the causes of OSCCs in the eye region of cattle. Methods: Sixty tumoral masses taken form 60 cattle with proliferation in the eye region that were collected between the years 2012-2022 were used. These cases were admitted to our department for routine diagnosis. The tissues were diagnosed as OSCC using histopathological methods. The presence of bovine papillomavirus (BPV), one of the causative factors, was investigated using immunohistochemical and polymerase chain reaction (PCR).

Results: Macroscopically masses were nodular or cauliflower-like and fragile and had hemorrhagic surfaces. Considering the keratin pearls, tumoral islands, and squamous differentiation, 20 out of 60 cases were classified as well, 20 as moderately, and 20 as poorly-differentiated OSCCs. 47 of the 60 cases were BPV positive using immunohistochemical methods. However, BPV nucleic acid was detected in only two cases with PCR. Only one of the cases could be sequenced. After phylogenetic analysis, virus strain was identified as BPV-1.

Conclusion: Our results indicated that papillomaviruses can contribute to the development of OSCCs, in both precursor lesions and also advanced stage OSCCs. We found that BPV-1 has a possible causative role; however, more studies are needed to investigate the role of other viral agents and their interaction with secondary factors.

Keywords: Bovine papillomavirus; Immunohistochemistry; Ocular squamous cell carcinoma; PCR.

Fig. 1

Solitary tumoral mass covering entire left eye. The lesion has cauliflower-like appearance, firm consistency, 8-9 cm diameter with hemorrhagic surface. Photographed from different angles (A-B)

Fig. 2

Images of the cases which are tested positive with PCR for BPV. (A-B) Poorly differentiated OSCC, mitotic figures (arrows) and dyskeratotic cells (dc) (H&E staining), (C) Poorly differentiated OSCC, BPV positive reactions in the cytoplasm of tumoral cells (arrows) in pleomorphic areas (IHC method), (D-E) Well differentiated OSCC, keratin pearl (KP), dyskeratotic cell (dc), and mitotic figure (arrow) (H&E staining), and (F) Severe intracytoplasmic BPV immunoreactivity in the cytoplasm of cells in tumoral islands (arrows) and inflammatory cell infiltration (arrowheads) around these islands

Fig. 3

Images of the cases which tested negative for BPV nucleic acid with PCR. BPV antibody is used for immunohistochemistry with chromogen substrate DAB (Hematoxylin staining). (A) Well-differentiated OSCC, BPV positive reactions in tumoral cells (arrows) and inflammatory cells (arrowheads), (B) Moderately differentiated OSCC, in the cytoplasm of neoplastic cells forming tumor islands (arrows), BPV immune-positive expressions, and (C) Poorly differentiated OSCC, severe intracytoplasmic (arrows) BPV, immunoreactivity in tumoral cells, especially in the periphery of tumoral islands

Fig. 4

Phylogenetic tree constructed using neighbor-joining (NJ) method using Kimura-Two parameter and 1000 bootstraps (MEGA 7.0) comparing our sequence with other reference papillomaviruses. Our sequence is indicated with black square (OK157506)

Fig. 5

Agarose gel image of PCR amplification with FAP59/FAP64 primer pair. Lane a: Positive control, Lane b: Sample, and Lane c: 100 bp (Thermo Scientific) DNA marker