Papillomaviruses are small, circular, double-stranded DNA viruses that infect a wide range of vertebrates, from humans and cats to horses and fish. In dogs, infections are usually subclinical, but certain viral types have been linked to hyperplastic lesions and, more rarely, to cancer. Thirty-three distinct canine papillomavirus types have been catalogued to date, organized into three genera: Lambdapapillomavirus, Taupapillomavirus, and Chipapillomavirus. These viruses are associated with a spectrum of epithelial disease, including oral and cutaneous papillomatosis, pigmented plaques, and squamous cell carcinomas.

Pigmented plaques in dogs are generally small, flat to slightly raised, darkly pigmented lesions that remain localized, most often affecting breeds such as pugs and miniature schnauzers. In a subset of cases, however, the plaques spread widely across the skin, and malignant transformation into squamous cell carcinoma has occasionally been documented. The mechanisms driving this progression remain incompletely understood, although the presence of viral oncogenes and host factors are thought to contribute. Until now, documented papillomavirus co-infections in dogs had been limited to interactions between CPV1 and CPV2, members of the Lambdapapillomavirus and Taupapillomavirus genera.

The patient in this report was referred to a veterinary dermatology service in Porto Alegre, Brazil, with cutaneous lesions that had been slowly progressing for five years. Prior treatments, including oral and injectable corticosteroids, monthly afoxolaner, and topical moisturizers, had met with limited success. Blood work was unremarkable and the dog was otherwise healthy. Physical examination revealed multiple pigmented plaques with black and yellow coloration on the darker skin of the trunk and limbs, erythematous crusted plaques on the lighter skin of the abdomen and neck, generalized comedones, and scaling. Several ulcerated lesions were also present, including a large one on the right side of the neck and two smaller ones on the flanks.

Under general anesthesia, the dog underwent skin biopsy. Histopathological examination of the pigmented plaques revealed an endophytic scalloped proliferation of spinous layer keratinocytes with hypergranulosis, hyperpigmentation, and orthokeratosis, consistent with canine pigmented viral plaque. The ulcerated neck lesion, by contrast, showed a nodular infiltrative transformation with large polygonal tumor cells forming trabeculae and keratin pearls, marked cellular atypia, and aberrant mitotic figures, supporting a diagnosis of moderately well-differentiated squamous cell carcinoma. A third sample from a flank lesion was classified as carcinoma in situ. Although the team could not confirm whether the carcinoma arose directly from a pre-existing plaque, it developed in a previously affected region of skin.

To characterize the viruses, the researchers turned to multiply primed rolling-circle amplification coupled with high-throughput sequencing, an approach well suited to circular viral genomes. The method enriches circular DNA and, unlike conventional PCR with degenerate primers, can recover divergent or co-infecting papillomavirus types without bias. Sequencing on an Illumina MiSeq platform generated more than 4.3 million paired-end high-quality reads. Of these, roughly 3.9 million were classified as CPV9 and about 409,000 as CPV8, and de novo assembly yielded two complete, distinct circular genomes, named CPV9_LMM and CPV8_LMM, measuring 8,396 and 7,953 base pairs respectively.

The two genomes showed average sequencing depths of 20,509-fold and 4,412-fold, ensuring reliable identification of every genomic region. Each carried the typical papillomavirus gene set, including the early genes involved in replication and cell transformation and the late genes encoding structural proteins for virion assembly, along with a non-coding upstream regulatory region controlling gene expression. Nucleotide identity with reference genomes reached 99.64 percent for CPV9 and 99.85 percent for CPV8, and phylogenetic reconstruction based on the L1 gene reinforced the classification of both viruses within the Chipapillomavirus genus. A negative control processed alongside the samples yielded no papillomavirus reads, and no other positive samples were handled simultaneously, supporting the authenticity of the dual detection.

DNA extracted from the paraffin-embedded carcinoma was then tested by conventional PCR targeting a partial L1 gene segment, followed by bidirectional Sanger sequencing. The carcinoma tested positive for viral DNA, and the sequence matched CPV8 with 100 percent nucleotide identity to the genome recovered from the pigmented plaques. The absence of CPV9 in this assay likely reflects primer complementarity bias, since consensus primers can preferentially amplify one viral genome over another in mixed infections. The researchers note that high-throughput sequencing, by contrast, enabled unbiased recovery of both complete genomes, underscoring its superior sensitivity for detecting co-infections.

Therapeutically, the ulcerated lesions were surgically excised, and the remaining plaques were treated with topical imiquimod 5 percent cream applied once daily. After 21 days the dog showed an excellent clinical response, with complete resolution or flattening of most plaques, although adverse effects included skin depigmentation and a focal inflammatory burn-like reaction on one flank that resolved when treatment was paused. Imiquimod was restarted at a reduced frequency of twice weekly. Fourteen months after the initial presentation, most lesions remained well controlled, but a new ulcerated lesion appeared on the right side of the body, and histopathology confirmed carcinoma in situ. That later lesion could not be examined molecularly.

The authors emphasize that the malignant potential of these canine papillomaviruses remains hypothetical, and that the CPV DNA detected in the carcinoma could conceivably originate from superficial epithelium or adjacent viral plaques rather than from the tumor cells themselves. A limitation of the study is the absence of tissue localization methods such as in situ hybridization or immunohistochemistry, which could have strengthened the evidence. Nevertheless, the consistent detection of the same CPV8 strain in both the plaque and the carcinoma, using different methodologies, supports an association between viral infection and neoplastic progression in this case. The finding that both co-infecting viruses encode E6 and E7, oncogenes known in papillomaviruses to drive continuous replication of infected epithelial cells and disrupt critical cellular regulatory pathways, raises the hypothesis that co-infections within the Chipapillomavirus genus could contribute to squamous cell carcinoma development in some canine patients. Notably, this transformation occurred in a dog with no clinical or laboratory evidence of immunodeficiency, reinforcing the need to reconsider the oncogenic potential of canine papillomaviruses in naturally occurring disease and to investigate the host-virus interactions that may tip pigmented plaques toward cancer.

Subject of Research: Co-infection of canine papillomavirus types 8 and 9 in pigmented viral plaques concurrent with squamous cell carcinoma in a dog

Article Title: Case Report: Canine papillomavirus type 8 and 9 co-infection in pigmented viral plaques concurrent with squamous cell carcinoma in a Chinese crested dog

Article References: Baretta, L. T., Saueressig, A., Pereira, E. D. L., Pereira, V. M. A. G., Schuch, M., Filippi, M., Sita, A., Schons, J. G., Demoliner, M., de Camargo, L. J., Budaszewski, R. F., Weber, M. N., de Almeida, P. R., Daudt, C., da Silva, F. C., Santos, K. H. S., Canal, C. W., Spilki, F. R., Trevizan, L., & da Silva, M. S. (2026). Case Report: Canine papillomavirus type 8 and 9 co-infection in pigmented viral plaques concurrent with squamous cell carcinoma in a Chinese crested dog. Frontiers in Virology, 6, Article 1832666. https://doi.org/10.3389/fviro.2026.1832666

Image Credits: AI Generated

DOI: 10.3389/fviro.2026.1832666

Keywords: canine papillomavirus, CPV8, CPV9, co-infection, pigmented viral plaques, squamous cell carcinoma, E6 oncogene, E7 oncogene, rolling-circle amplification, high-throughput sequencing, veterinary dermatology, oncogenesis