Deer Papillomavirus Research Articles

Papillomavirus infection of roe deer in the Czech Republic and fibropapilloma-associated levels of metallothionein, zinc, and oxidative stress

The present study aimed at the aetiological diagnosis of skin tumours of roe deer (Capreolus capreolus) in the Czech Republic. A total of 33 roe deer specimens showing skin masses were sampled for histopathology, virus detection and identification using polymerase chain reaction (PCR) and deoxyribonucleic acid sequence (DNA) analysis, and to investigate fibropapilloma-associated levels of metallothionein, zinc and oxidative stress in 2012 and 2013. Ticks (Ixodes ricinus) and deer keds (Lipoptena cervi) were also collected and pooled from sampled game specimens. Ticks found on dogs in hunting grounds under study were pooled into another sample. Skin tumours, ranging from 2 to 10 per inspected specimen and 2 to 5 cm in size, were classified as fibropapillomas by histopathology. All 33 viral-infection-suspected skin samples from roe deer in South Moravia and South Bohemia were PCR positive. The nucleotide sequences of PCR products were 100% homologous to the Western roe deer papillomavirus 1 isolate CcPV-1. Ticks and deer keds from positive roe deer and ticks from dogs were PCR and DNA sequence positive for the roe deer papillomavirus. Viral DNA was also demonstrated in one blood sample from a roe deer female. Differences in metallothionein, zinc, taurine and electrochemical index among samples from lesions and normal skin of affected roe deer and negative controls from papillomavirus-non-infected animals were non-significant. While we have demonstrated circulation of specific roe deer papillomavirus in the Czech Republic that results in multiple fibropapillomatous skin tumours, many ecological and epidemiological issues of this wildlife disease still remain unanswered.

From Amino Acids Profile to Protein Identification: Searching for Differences in Roe Deer Papilloma

Papillomaviruses, small non-enveloped DNA viruses, are considered as the cause of a number of cancers, such as a cervical, skin, anal, and penile cancers. Roe deer papillomavirus infection can be easily characterized by typical warts on the skin. The aim of this study was to characterize differences of amino acid and protein composition in healthy and tumour tissue of roe deer using methods including ion-exchange chromatography, SDS-PAGE, two-dimensional gel electrophoresis and MALDI-TOF MS. The obtained data were statistically evaluated. Cluster analysis of all variables showed biggest differences in proline in control and papilloma samples. Further, we aimed at electrophoretic analysis of the samples. Discriminant analysis projections to latent structures (Projection to the latent structure Discrimination analysis, syn. Discrimination Partial least squares analysis, PLS DA) was used to evaluate the obtained results. Examining model PLS DA, there was found that the greatest effect on whether the individual is classified as healthy, or with papilloma, is the latent variable that best correlates with the molecular mass from 108.8 to 128.1 kDa and from 231.2 to 290.4 kDa in the original signal. Finally, using mass spectrometry three proteins, pyridoxal kinase, myoblast determination protein and leucine zipper transcription factor-like protein 1, most likely related to cancer were found.

Papillomavirus-associated fibropapillomas of red deer ( Cervus elaphus )

Oval, firm, cutaneous tumours with a rough, hairless, pigmented surface, exhibiting a moderately pronounced papillary structure were detected on the abdominal skin of two young red deer ( Cervus elaphus ). One animal was shot in Lower Austria in 2004, the other at a deer farm in Hungary in 2007. Histological examination of both samples classified the tumours as fibropapillomas, showing marked proliferation of fibroblasts and connective tissue, accompanied by hyperkeratosis, parakeratosis and acanthosis of the overlaying epidermis, and occasional foci of inflammation. The distribution of cytokeratin and vimentin was characterised in the lesion. The presence of papillomavirus (PV) antigen was demonstrated by immunohistochemistry in both cases. Papillomavirus-specific DNA was successfully amplified by PCR from one sample. The obtained partial nucleotide sequence of the L2 ORF exhibited the highest critical identity values with the homologous regions of Delta-papillomaviruses, especially the Roe deer papillomavirus (93%). Phylogenetic analysis of the partial L2 ORF sequence alignment of 10 papillomaviruses by both neighbour-joining and maximum parsimony method confirmed that the Red deer PV is very closely related to the Western roe deer papillomavirus (CcPV1).

Endemic papillomavirus infection of roe deer ( Capreolus capreolus)

Roe deer papillomavirus (CcPV1) infection has been identified as an endemic disease in roe deer populations of the Carpathian basin in Central Europe (Hungary, Austria and Croatia). The disease is characterised by easily recognizable skin tumours similar to deer papillomavirus infection of North American deer species. In 2006, a questionnaire study was conducted among all Hungarian game management units (GMUs) in order to assess the distribution of the disease and its major epidemiological features. Categorical information was collected about disease occurrence, trend and frequency of detection, on primarily affected age classes in both sexes, and association of lesions with mortality. Replies were received from 539 GMUs representing 50.9% of total GMU territory and disease presence was reported by 295 (54.7%) GMUs. Older age classes of both sexes were found to be more affected. Association of various environmental factors with disease occurrence was evaluated and data were collected on the occurrence of similar skin lesions in other European countries. Pathological features of CcPV1 infection were described and the localisation of both CcPV1 antigen and DNA was characterised by immunohistochemistry and in situ DNA hybridisation in skin lesions. Virus presence was also demonstrated by PCR and PCR product sequencing.

Characterisation of the first complete genome sequence of the roe deer (Capreolus capreolus) papillomavirus

The complete genomic DNA of a novel roe deer (Capreolus capreolus) papillomavirus (CcPV1) was amplified and sequenced from fibropapillomatous skin lesions of a Hungarian roe deer. Viral DNA was detected by a pair of degenerate primers and the remaining genomic sequence was amplified by a long-template high-fidelity PCR and sequenced. The CcPV1 genome was 8032bp long and contained open reading frames (ORFs) typical for Delta-papillomaviruses (E6, E7, E1, E2, E4, E5, E9, L2, and L1) and a 799bp long untranslated regulatory region (URR). Phylogenetic analysis based on the 3861bp long concatenated sequence of the E1–E2–L2–L1 ORFs and on separate alignments of all major ORFs using both neighbour-joining and maximum parsimony methods placed CcPV1 on a distinct branch between Ovine papillomavirus 1 and the other deer papillomaviruses within the Delta-papillomavirus genus, although pairwise nucleotide alignments of L1 ORF sequences determined highest identities with European Elk Papillomavirus (71.2%) and Reindeer Papillomavirus (70.3%).

Detection of papillomavirus-DNA in mesenchymal tumour cells and not in the hyperplastic epithelium of feline sarcoids.

We examined 12 formalin-fixed paraffin-embedded feline skin tumours which had the histopathological features of fibropapillomas for the presence of papillomavirus (PV) DNA using touchdown polymerase chain reaction (PCR), DNA sequencing and nonradioactive in situ hybridization. Nine of the tumours contained a 102-bp PCR product demonstrated using consensus PV primers that amplify a portion of the L1 gene. The nucleotide sequences are closely related, but not identical to that of ovine PV type 2, rabbit oral PV and reindeer PV. The deduced amino acid sequences had strong homologies with the major capsid protein L1 of deer PV, bovine papillomavirus (BPV) 1 and BPV 2, and European elk PV. Although PV antigens were not detected in any of the tumours by immunohistochemistry, PV DNA was demonstrated in individual mesenchymal cells or cell nests of 4/12 tumours by in situ hybridization. A nonproductive infection of mesenchymal fibroblast-like tumour cells with a papillomavirus would explain the lack of PV antigen expression and the absence of PV DNA in the hyperplastic epithelium. Because these tumours and their pathogenesis are similar to equine sarcoids, we suggest that they should be reclassified as 'feline sarcoids' instead of fibropapillomas.

Identification of Conformational Epitopes of the BPV-1 Capsid Recognized by Competitive Inhibition of Sera from Infected or Immunized Animals

Conformational papillomavirus (PV) capsid epitopes are the major targets of neutralizing antibodies. BPV1 virion surface epitopes were analyzed for type-specificity, induction of neutralization, and topographical location with antibodies from selected humans, and BPV1 or -2 immunized or infected cattle, rabbits and mice. Rabbit sera produced against deer PV (DPV) or BPV2 and human sera reacted with intact BPV1 in ELISA, indicating the presence of cross-reactive conformational epitopes on the surface of BPV1 virions. However, competitive binding assays performed on intact BPV1 capsids using unlabeled and biotin-conjugated antibodies revealed that the conformational epitopes of BPV1 recognized by rabbit, bovine and human antibodies were topographically different. Only hyperimmune sera to intact BPV1, BPV2 or DPV neutralized BPV1-induced focus formation of C127 cells with the highest neutralizing titers observed for bovine and rabbit antibodies produced against intact BPV1. Our study showed the complexity of conformational capsid epitopes of BPV1 and their importance for inducing a protective humoral immune response in animals inoculated or immunized with BPV1.

Reindeer papillomavirus transforming properties correlate with a highly conserved E5 region.

A papillomavirus was isolated from the epithelial layer of a cutaneous fibropapilloma on a Swedish reindeer (Rangifer tarandus). Reindeer papillomavirus (RPV) is morphologically indistinguishable from other papillomaviruses, but the restriction enzyme cleavage pattern of its genome is different. No sequence homology was detected between RPV DNA and the DNAs of bovine papillomavirus type 1 (BPV-1) and avian papillomavirus when hybridization was performed under stringent conditions. However, the RPV genome hybridized to the genome of the European elk papillomavirus and the deer papillomavirus under stringent conditions. A physical map of the RPV genome was constructed, and selected regions of the genome, covering the open translational reading frame (ORF) E5 and part of the E1 and L1 ORFs, were studied by nucleotide sequence analysis. The results made it possible to align the RPV genome with the genome of BPV-1. The E5 ORF of RPV has the potential to encode a 44-amino-acid, exceptionally hydrophobic polypeptide which is very similar to the E5 polypeptides of BPV-1 and deer and European elk papillomaviruses. RPV is oncogenic for hamsters and transforms C127 mouse cells in vitro. Several virus-specific mRNAs were detected in RPV-transformed C127 cells.

Topographical and Conformational Epitopes of Bovine Papillomavirus Type 1 Defined by Monoclonal Antibodies<xref ref-type="fn" rid="FN2">2</xref>

Monoclonal antibodies (MAbs) were generated against sodium dodecyl sulfate-disrupted bovine papillomavirus type 1 (BPV-1). When screened by enzyme-linked immunosorbent assay (ELISA) on intact and disrupted BPV-1, -2, and deer papillomavirus, three patterns of reactivity were defined: reactivity only with intact virus, with both intact and disrupted virus, and only with disrupted virus. On the basis of ELISA results, the topographical location and requirement for conformation for immunoreactivity of epitopes was defined as external conformational, external linear, and internal linear. Cross-reactivity of MAbs with other papillomavirus types was analyzed by immunofluorescence on warts from different species. Type-specific, BPV-1 and/or -2 cross-reactive, broadly cross-reactive, and genus-specific MAbs were identified. MAb reactivity with structural polypeptides of BPV-1 was analyzed by Western blot. MAbs reactive with epitopes defined as conformational by ELISA did not react in Western blot. All MAbs reactive in Western blot reacted with the major capsid protein (MCP) [55 kilodalton (kDa)], demonstrating that the MCP carries both type-specific and cross-reactive epitopes. Most MAbs reactive with the MCP were cross-reactive with structural polypeptides of 48 and 96 kDa, demonstrating the immunologic relatedness of these three polypeptides.

Organization and expression of the transforming region from the European elk papillomavirus (EEPV)

The nucleotide sequence of the early (transforming) region from the European elk papillomavirus (EEPV) double-stranded DNA has been determined together with flanking regions. The established sequence, which is 5732 bp long, shows that the genome of EEPV is closely related to the previously sequenced bovine papillomavirus type 1 (BPV-1) and deer papillomavirus (DPV) genomes. Seven open reading frames (ORFs), designated E1–E7, were identified in similar positions as in the BPV-1 genome. The E1 and E5 regions were best conserved. The strong homology between the E5 ORFs of EEPV, BPV-1 and DPV is interesting in the light of the recent proposal that these ORFs encode a major transforming function (Schiller et al., 1986; DiMaio et al., 1986). A set of mRNAs, comprising six size classes, were identified in EEPV-transformed cells. At least two different promoters appear to control EEPV transcription in transformed cells.

A cutaneous fibropapilloma from a red deer (Cervus elaphus) associated with a papillomavirus.

A cutaneous fibropapilloma was found on a Scottish red deer (Cervus elaphus), and a papillomavirus was isolated from it. The virus appeared to be related to bovine papillomavirus type 1 (BPV1) or type 2 (BPV2) because: (i) it cross-reacted in peroxidase-antiperoxidase tests with antisera raised against these virions; (ii) BPV1 and BPV2 DNAs cross-hybridized to the red deer papillomavirus in situ; and (iii) BPV1 and/or BPV2 DNA cross-hybridized to the red deer papillomavirus DNA on Southern blots under conditions of high stringency. These tests also revealed a unique restriction enzyme cleavage pattern for the red deer papillomavirus DNA.