Felis Catus Papillomavirus Research Articles

Felis catus papillomavirus type-2 but not type-1 is detectable and transcriptionally active in the blood of healthy cats.

Papillomaviruses (PVs) are small DNA viruses that induce benign and/or malignant epithelial tumours in different species, including the domestic cat (Felis catus). To date, five F.catus papillomavirus genotypes have been identified (FcaPV-1 to FcaPV-5). FcaPV-1 is associated with skin and oral benign lesions, while FcaPV-2 infection is widely associated with feline squamous cell carcinomas. Several human and animal PVs have been found in body fluids such as peripheral blood; however, the presence of FcaPVs in non-epithelial tissues has not previously been investigated. The aim of this study was to assess the presence and gene expression of FcaPV-1 and FcaPV-2 in the blood of healthy cats. We detected FcaPV-2 DNA in 26 of 103 (25%) blood samples. Importantly, FcaPV-2L1, E2, E6 and E7 genes were found to be expressed in 3 (25%), 11 (92%), 6 (50%) and 5 (42%) of the samples available for mRNA analysis, respectively. FcaPV-1 was not detected in any of the blood samples analysed here. The data obtained in this work suggest active and eventually productive infection of FcaPV-2 in the blood of healthy cats, implying a possible role in intra-individual spreading as well as in vertical and horizontal transmission.

A study of multiple Felis catus papillomavirus types (1, 2, 3, 4) in cat skin lesions in Italy by quantitative PCR.

Objectives The aim of the study was to investigate, by quantitative PCR (qPCR), the presence of papillomavirus in feline viral plaques (VPs), Bowenoid in situ carcinoma (BISC), squamous cell carcinoma (SCC) and actinic keratosis (AK). Methods Twenty-nine cases with previously established diagnoses of feline VPs, BISC, invasive SCC and AK were selected from a dermatopathological database. A critical re-evaluation of diagnosis was performed by defining clear criteria toward carcinomatous vs non-carcinomatous, in situ vs invasive (if carcinomatous) and viral vs actinic. Cases were evaluated for p16 immunolocalisation. The presence of the target viral genes for Felis catus papillomavirus (FcaPV)-1, FcaPV-2, FcaPV-3 and FcaPV-4 was determined by qPCR. The data generated ΔΔCq values, which represent a normalised measure of DNA viral quantity. Samples with a positive ΔΔCq value were submitted to sequence analysis. Results Four VPs, 19 BISCs, four SCCs and one case of AK were included. By ΔΔCq analysis we found that all VPs were positive for FcaPV-1 or FcaPV-2; eight BISCs were positive for FcaPV-1, FcaPV-2 and FcaPV-4. FcaPV-2 was the most prevalent among the group of VPs and BISCs. Conclusions and relevance Using the ΔΔCq method we report the first evidence of FcaPV-1, FcaPV-2 and FcaPV-4 in Italy. FcaPV-2 was the most frequently detected; to a lesser extent, FcaPV-1 and FcaPV-4 were detected in the examined samples. FcaPV-3 was never associated with viral-induced lesions by ΔΔCq investigation. Compared with conventional PCR the ΔΔCq method has the advantage of establishing a possible role of the virus in the outcome of infection.

Genomic characterisation of Felis catus papillomavirus type 5 with proposed classification within a new papillomavirus genus

Four Felis catus papillomavirus (FcaPV) types have been fully sequenced from domestic cats. Of these, FcaPV-2 and -3 are thought to cause feline viral plaques and Bowenoid in situ carcinomas. Two short sequences of DNA from a previously unreported PV type were amplified from a feline viral plaque using consensus PCR primers. DNA was then extracted from a swab of the lesion and two sets of ‘outward facing’ primers were designed using the short sequences to amplify the entire 7600bp genome of the novel PV. The PV was designated FcaPV-5 and contained putative coding regions that were predicted to produce five early proteins and two late ones. The ORF L1 showed over 65% similarity to that of FcaPV-3 and -4. Assignment to a genus was difficult as the PV was over 60% similar to PV types from 4 different genera. However, due to the ORF L1 similarity of FcaPV-3, -4, and -5, the shared host species of all three PVs, and the similar lesions associated with FcaPV-3 and -5, it is proposed all three PVs are classified within a new genus. FcaPV-5 is the third PV type to be associated with feline viral plaques. The plaque that contained FcaPV-5 showed unusual histological features including hyperplasia and PV-induced cell changes in sebaceous glands and deep within hair follicles. While additional study of further lesions of this type is required, it is possible that FcaPV-5 may be able to infect a broader range of cells than other PV types.

Multiple viral plaques with sebaceous differentiation associated with an unclassified papillomavirus type in a cat

CASE HISTORY AND CLINICAL FINDINGSA 15-year-old neutered male domestic short-haired cat was presented due to multiple 0.5–2 cm-diameter crusting plaques in the left preauricular region, over the bridge of nose, and in the right periocular region. The plaques did not appear to cause discomfort.HISTOPATHOLOGICAL FINDINGSBiopsy samples of four plaques were examined histologically. Three plaques consisted of well-demarcated foci of mild epidermal hyperplasia overlying markedly hyperplastic sebaceous glands. Approximately 60% of the hyperplastic cells contained a large cytoplasmic vacuole that ranged from being clear to containing prominent grey-blue fibrillar material. The fourth plaque was composed solely of epidermal hyperplasia, consistent with previous descriptions of feline viral plaques.MOLECULAR BIOLOGYPapillomavirus DNA was amplified from all four plaques using PCR. A single DNA sequence was amplified from the plaques with sebaceous differentiation. This sequence was identical to the FdPV-MY sequence previously suggested to be from a putative unclassified papillomavirus type. Felis catus papillomavirus type 2 sequences were amplified from the plaque typical of feline viral plaques. Immunohistochemistry to detect p16CDKN2A protein (p16) showed marked immunostaining throughout the hyperplastic epidermis and adnexal structures within the plaques with sebaceous differentiation.DIAGNOSISMultiple feline viral plaques with variable sebaceous differentiation.CLINICAL RELEVANCEFeline viral plaques with sebaceous differentiation have not been previously reported in cats. The presence of unique cell changes within these lesions, the detection of an unclassified papillomavirus type, and the p16 immunostaining within these plaques suggest that they may have been caused by the papillomavirus that contains the FdPV-MY sequence.

Felis catus papillomavirus type 2 E6 oncogene enhances mitogen-activated protein kinases and Akt activation but not EGFR expression in an in vitro feline model of viral pathogenesis

A possible causative role of Felis catus papillomavirus type 2 (FcaPV2) in the development of feline oral and cutaneous squamous cell carcinomas (SCC) has been recently suggested by demonstrating viral gene expression in vivo and transforming properties by its putative oncogenes E6 and E7 in vitro. The activated molecules MEK (pMEK), ERK (pERK) and Akt (pAkt) are signaling transduction effectors regulating cell proliferation and inhibition of apoptosis, which are critical steps towards tumour formation. Here, we show by Western blotting (WB) that expression of FcaPV2 E6 in feline epithelial cells enhances pMEK, pERK and pAkt levels compared to control cells. Additionally, we demonstrated by real-time quantitative PCR on epidermal growth factor receptor (EGFR) transcripts and WB that activation of these signaling routes is independent from EGFR differential gene expression, total protein levels or phosphorylation, unlike in human papillomavirus associated tumours. This study contributes to define the molecular scenario underlying FcaPV2-triggered pathogenesis of feline SCC.

Transforming properties of Felis catus papillomavirus type 2 E6 and E7 putative oncogenes in vitro and their transcriptional activity in feline squamous cell carcinoma in vivo

Felis catus papillomavirus type 2 (FcaPV2) DNA is found in feline cutaneous squamous cell carcinomas (SCCs); however, its biological properties are still uncharacterized. In this study, we successfully expressed FcaPV2 E6 and E7 putative oncogenes in feline epithelial cells and demonstrated that FcaPV2 E6 binds to p53, impairing its protein level. In addition, E6 and E7 inhibited ultraviolet B (UVB)-triggered accumulation of p53, p21 and pro-apoptotic markers such as Cleaved Caspase3, Bax and Bak, suggesting a synergistic action of the virus with UV exposure in tumour pathogenesis. Furthermore, FcaPV2 E7 bound to feline pRb and impaired pRb levels, resulting in upregulation of the downstream pro-proliferative genes Cyclin A and Cdc2. Importantly, we demonstrated mRNA expression of FcaPV2 E2, E6 and E7 in feline SCC samples, strengthening the hypothesis of a causative role in the development of feline SCC.

Frequent detection of transcriptionally active Felis catus papillomavirus 2 in feline cutaneous squamous cell carcinomas.

Felis catus papillomavirus 2 (FcaPV-2) causes premalignant skin lesions in cats and has also been found in a proportion of cutaneous squamous cell carcinomas (SCCs) - a common and potentially fatal cancer of cats. Whilst this could suggest a role of the virus in cancer development, FcaPV-2 has also been detected in skin swabs of normal cats, making it difficult to discern whether the papillomavirus is causing the cancer or merely an 'innocent bystander'. To distinguish between these two possibilities, real-time PCR was used to determine the viral copy number and the transcriptional activity of FcaPV-2 infections present in 70 formalin-fixed paraffin-embedded skin lesions including 10 papillomavirus-induced premalignant lesions and 60 SCCs. FcaPV-2 gene expression was found in 21 of 60 (35 %) SCCs, all 10 premalignant lesions and none of 10 normal skin samples. The results showed two distinct subsets of SCCs. The majority of the SCCs had low copy numbers of FcaPV-2 DNA (mean of 17 copies per copy of reference gene DNA) and no FcaPV-2 gene expression, suggesting the virus was an incidental finding. In contrast, 20 SCCs had detectable FcaPV-2 E6/E7 gene expression and very high copy numbers of FcaPV-2 DNA, with a mean of 32 930 copies per copy of reference gene DNA. The relative quantity of E6/E7 gene expression and the viral copy number in this group were similar to those found in the papillomavirus-induced premalignant lesions, suggesting that FcaPV-2 may play a role in the development of a subset of feline cutaneous SCCs.

Felis catus papillomavirus types 1 and 4 are rarely present in neoplastic and inflammatory oral lesions of cats

Oral squamous cell carcinomas (OSCCs) are common feline cancers. Why OSCCs are so common in cats is unknown; however, 25% of human OSCCs are caused by papillomaviruses (PVs). Two feline oral PVs (FcaPV-1 and 4) are recognized. As PVs are highly host and location specific, if PVs do cause feline OSCCs, FcaPV-1 and 4 are the most likely etiological agents. PCR primers specific for FcaPV-1 amplified DNA from 1 of 36 feline OSCCs and 1 of 16 inflammatory oral lesions. No DNA was amplified by primers specific for FcaPV-4. PV DNA was not amplified from any additional sample using consensus primers. No PV cytopathology was visible in the OSCC that contained FcaPV-1 DNA, but viral cytopathology was present in a focus of epithelial hyperplasia in the non-neoplastic sample. This study does not support a PV etiology of feline OSCCs, but shows that FcaPV-1 can asymptomatically infect the mouth of cats.

The use of quantitative PCR to detect Felis catus papillomavirus type 2 DNA from a high proportion of queens and their kittens

Squamous cell carcinomas are common feline skin cancers that have been associated with infection with Felis catus papillomavirus type 2 (FcaPV-2). Currently, little is known about the epidemiology of FcaPV-2 infection. The aim of this study was to develop a real-time PCR assay to quantify FcaPV-2 DNA in plucked hairs and skin swabs from 11 healthy breeding queens and their kittens. Samples were taken prior to kittening and then 2, 7 and 28 days after kittening to determine the age at which the kittens were first exposed to the virus. FcaPV-2 DNA was amplified from all of the queens and from 91% of the kittens at 2 days of age. There was a wide range in the quantity of FcaPV-2 DNA detected, from 1 to 92,520 copies per swab, and from 0.01 to 234 copies per copy of reference gene DNA in the hair plucks. The quantity of FcaPV-2 DNA detected in samples collected from the kittens was strongly correlated to that of their respective queens and the mean viral DNA load was similar for cats within a household but varied significantly between households. This is the first time that quantitative PCR has been used to detect FcaPV-2 DNA and the results suggest that the virus is ubiquitous but there is a wide variation of viral DNA loads. Kittens appear to be exposed to FcaPV-2 early in life, presumably from direct contact with their queen. These results are important when determining if FcaPV-2 infection of cats is preventable.

Genomic characterisation of Felis catus papillomavirus 4, a novel papillomavirus detected in the oral cavity of a domestic cat

Three papillomaviruses (PVs) from the domestic cat have been fully sequenced so far including Felis domesticus PV-1 (FdPV-1), FdPV-2, and a recently described Felis catus PV-3 (FcaPV-4). In the current article, we describe the full genomic sequence of a fourth PV from the domestic cat. This PV was amplified from the oral cavity of a cat with severe gingivitis. However, the aetiological involvement of FcaPV-4 in development of lesions observed in this cat remains uncertain. The complete genome of the novel virus comprised 7,616 bp and was predicted to encode five early (E1, E2, E4, E6 and E7) and two late (L1 and L2) genes, with the organisation typical for PVs. The L1 showed 65.1 % nucleotide sequence identity to L1 of FcaPV-3 and approximately 60 % identity to L1 of canine tau-papillomaviruses CPV-2 and CPV-7. The novel virus clustered with FcaPV-3, CPV-2 and CPV-7 on a phylogenetic tree constructed from a concatenated alignment of 3,013 bp from E1, E2, L1 and L2. Based on the genomic and phylogenetic data, we propose that the novel virus is classified as a distinct species within the same genus as FcaPV-3. We also propose that both viruses are classified within the genus Taupapillomavirus, although this classification may need to be re-visited after more tau-PV genomes become available.