Papillomavirus infection of aged Persian cats

Papillomavirus infection was confirmed in 2 Persian cats with sessile hyperkeratotic skin lesions. Skin lesions were not typical papillomas as found in other species. Papillomavirus were demonstrated in negative stain preparations of homogenized tissue and within nuclei of cells in the stratum granulosum. Papillomavirus group-specific antigens were detected within nuclei corresponding to those containing virions. Attempts to transmit this disease to other cats or propagate the virus in tissue cultures were unsuccessful. A 7.8-kilobase DNA molecule was present in low-stringency Southern blots using a bovine papillomavirus type 1 cloned DNA probe. In reverse Southern blots, the cat papillomavirus hybridized under conditions of low stringency with all papillomavirus genomes tested. Combined with limited restriction endonuclease restriction mapping, the above information indicates that the feline cutaneous papillomavirus is a unique virus type and thus expands the list of hosts known to be infected by papillomaviruses.     

Molecular characteristics of cutaneous papillomavirus from the canine pigmented epidermal nevus

To investigate the relation between the canine pigmented epidermal nevus (PEN) and cutaneous papillomavirus, we cloned and sequenced the L1 gene of papillomavirus from the canine pigmented epidermal nevus (PEN). Amplification of DNA sample with the L1 consensus primers yielded an expected fragment of approximately 450-bp. The nucleotide sequences of the fragment showed about 64% of sequence similarity to the L1 region of human papillomavirus isolate CP6108 and less than 57% sequence similarity to those of canine oral papillomavirus (COPV). In situ hybridization determined the presence of papillomavirus DNA mainly in the upper stratum granulosum of skin in this case. The results indicated that the canine cutaneous papillomavirus from the PEN lesion was genetically close to human papillomavirus rather than COPV.     

Canine inverted papillomas associated with DNA of four different papillomaviruses

Inverted papillomas are uncommon papillomavirus (PV)‐induced canine skin lesions. They consist of cup‐ to dome‐shaped dermal nodules with a central pore filled with keratin. Histologically they are characterized by endophytic projections of the epidermis extending into dermis. Cytopathic effects of PVs infection include the presence of clumped keratohyalin granules, koilocytes and intranuclear inclusion bodies. Different DNA hybridization studies carried out with a canine oral papillomavirus (COPV) probe suggested that a different PV than COPV might cause these lesions. Canine papillomavirus 2 (CPV2) was discovered a few years ago in inverted papillomas of immunosuppressed beagles. Two other cases, presenting with distinct clinical and histological features have also been described. This study was carried out on four dogs with clinical and histological signs of inverted papillomas. Molecular biological analyses confirmed that PV DNA was present in all four lesions but demonstrated that the sequences in each case were different. One corresponded to COPV, the second to CPV2, and the third and fourth to unknown PVs. These findings suggest that inverted papillomas are not caused by one single PV type. Similar observations have also been made in human medicine.     

Papillomas and carcinomas associated with a papillomavirus in European harvest mice (Micromys minutus)

Papillomaviruses, group-specific papillomavirus antigens, or extrachromosomal papillomavirus DNA were detected in cutaneous, mucocutaneous, and pulmonary tumors affecting a colony of European harvest mice (Micromys minutus). Skin lesions were classified as acanthomatous hyperplasia, epidermal inclusion cysts, squamous papillomas, inverted papillomas, trichoepitheliomas, and sebaceous carcinomas. Cutaneous horns (hyperkeratotic papillomas) were on mucocutaneous junctions of one animal. One mouse, with a cutaneous sebaceous carcinoma, had multiple pulmonary keratinaceous cysts. Papillomavirus antigens, detected by the avidin-biotin technique, were in 20 of 31 lesions tested. In contrast, by Southern blot hybridization all 28 lesions tested contained papillomavirus DNA. Papillomavirus DNA was demonstrated in two of ten benign cutaneous lesions by in situ hybridization.     

Cloacal papillomas in psittacines

Papilloma-like masses affecting the cloaca of 19 Amazons, 16 macaws, 3 parrots, 1 conure, and 1 parakeet were examined. Papillomatous lesions were characterized by proliferation of the lining epithelium on thin fibrovascular stalks. Carcinoma in situ was diagnosed in the cloaca of a macaw in addition to the other 16 macaws with papillomas. Papillomavirus group-specific antigens were not detected in any of the 41 lesions, using the peroxidase-antiperoxidase technique. The DNA extracts from 6 different frozen papillomas did not contain papillomavirus genomes detectable by Southern blot hybridization, using an African gray parrot cutaneous papillomavirus as a probe. Evidence of an infective agent was not found by electron microscopic examination of 8 of the papillomas. Inoculations of partially purified homogenates of a cloacal papilloma from a yellow-crowned Amazon did not induce lesion formation on cloacal mucosa of an adult yellow-crowned Amazon, green and yellow macaw, sulphur-crested cockatoo, or mollucan cockatoo.     

Cutaneous neoplasms in dogs associated with canine oral papillomavirus vaccine

A spectrum of proliferative cutaneous lesions occurred in 12 dogs at the injection site of live canine oral papillomavirus (COP) vaccine, suggesting a viral etiology for the masses. Lesions included epidermal hyperplasia, epidermal cysts, squamous papilloma, basal cell epithelioma, and squamous cell carcinoma. Peroxidase-antiperoxidase staining of tumor sections revealed nuclei which stained for group-specific papillomavirus antigen in five of 12 masses. Electron microscopic examination of tumor sections did not reveal virions. In transmission studies, macerated tumor tissue did not produce oral papillomas on the scarified mucosa of puppies; this procedure did protect the puppies from development of lesions when challenged with infectious papilloma material. These findings are evidence that COP can induce hyperplastic and neoplastic lesions in sites other than oral, pharyngeal, and ocular mucosa.     

Evaluation of papillomaviruses associated with cyclosporine-induced hyperplastic verrucous lesions in dogs

OBJECTIVE: To determine whether cyclosporine A-induced hyperplastic skin lesions of dogs were associated with papillomavirus infections. ANIMALS: 9 dogs that were treated with cyclosporine A and developed hyperplastic skin lesions. PROCEDURE: History and clinical and histopathologic data were collected. Paraffin-embedded skin biopsy specimens from hyperplastic skin lesions were immunostained for common papillomavirus genus-specific structural antigens by use of a polyclonal rabbit anti-bovine papillomavirus type 1 antiserum. Sections from each tissue block underwent DNA extraction, and polymerase chain reaction (PCR) assays were performed with several sets of primers to amplify a wide range of papillomavirus DNA from humans and other animals. RESULTS: In 7 of 9 dogs, there were more than 10 hyperplastic skin lesions that microscopically resembled those of psoriasiform lichenoid dermatosis. In those dogs, results of testing for papillomavirus via immunohistochemical analyses and PCR assays were negative. In the other 2 dogs, there were only 1 and 3 verrucous lesions, and in those dogs, histologic evaluation revealed koilocytes and nuclear viral inclusions that were immunoreactive for papillomavirus antigens. Papillomavirus DNA was amplified from both dogs. One of the sequences was characteristic for the canine oral papillomavirus, whereas the other had similarities with the recently described canine papillomavirus 2. CONCLUSIONS AND CLINICAL RELEVANCE: In dogs, hyperplastic skin lesions occasionally develop during treatment with cyclosporine A. Most of the lesions resemble those of psoriasiform lichenoid dermatosis, although papillomavirus can be detected in some instances.     

Investigation of parrot papillomavirus in cloacal and oral papillomas of psittacine birds

Cloacal and oral papillomas from 27 psittacine birds of various species were examined for the presence of parrot papillomavirus by DNA in situ hybridization, DNA in situ polymerase chain reaction, and nested polymerase chain reaction. Parrot papillomavirus was detected in one oral papilloma from an African grey parrot by all three techniques. In addition, rare basophilic intranuclear inclusions were observed by light microscopy in tissue sections of the oral papilloma from this parrot. The remaining lesions were negative for parrot papillomavirus DNA. This study suggests that parrot papillomavirus may be involved in the development of papillomas in African grey parrots, but apparently is not responsible for development of similar lesions in unrelated species of psittacine birds.     

The development of multiple cutaneous inverted papilloma following ovariohysterectomy in a dog

Abstract

CASE HISTORY: Ovariohysterectomy was performed on an adult Cavalier King Charles Spaniel. The skin that had been clipped for surgery was noticed to be erythematous 8 days later.

CLINICAL AND PATHOLOGICAL FINDINGS: Poorly defined patches containing multiple papules were visible bilaterally within the clipped skin. These became larger over the following 2 weeks, and samples were collected for histology. Seven days later, the lesions were multiple raised masses, up to 5 cm in diameter. Histology revealed numerous cup-shaped epidermal proliferations extending into the dermis. The presence of keratinocytes with increased quantities of blue-grey cytoplasm, and koilocytosis suggested papillomavirul infection; Canis familiaris papillomavirus (CfPV-2) DNA was amplified from two separate samples. Complete regression was observed 8 weeks after the lesions had been initially observed.

DIAGNOSIS: Multiple inverted papilloma confined to skin that had been clipped for surgery.

CLINICAL RELEVANCE: This is the first time that the development of canine cutaneous papillomas has been associated with surgery. The nature of the association between surgery and development of the papillomas is uncertain. However, it is possible that damage to superficial skin could promote the formation of papillomas. This is the first identification of CfPV-2 in New Zealand.     

Papillomavirus associated skin lesions in a cat seropositive for feline immunodeficiency virus.

A cat was presented with skin lesions consisting of slightly raised pigmented plaques, 2-7 mm in diameter with a rough slightly verrucous surface. Histologically these lesions were identified as papillomas. A papillomavirus infection was demonstrated: virus-like particles were present in the nuclei of cells within the lesions, and staining with an anti-bovine papillomavirus (BPV-1) antibody was obtained. An infection with feline immunodeficiency virus was diagnosed in this cat; this condition had probably enhanced the development of papillomas. This is the first report of a papillomavirus infection in a cat in Europe.     

Parapoxvirus infection in harbor seals (Phoca vitulina) from the German North Sea

In the summer of 2000, proliferative lesions of the skin and oral mucosa were observed in 26 young harbor seals (Phoca vitulina) from a rehabilitation center in Schleswig-Holstein, Germany. Verrucose, roundish nodules, approximately 1-2 cm in diameter, were presented in the oral cavity, especially on the tongue. Some animals developed similarly sized spherical dermal elevations with ulceration on flippers, chest, neck, and perineum. Necropsy of one animal showed multifocal, verrucose nodules in the oral cavity and a mild tonsillitis. Histologically, the nodules were characterized by ballooning degeneration of the outer parts of the spiny layer and stratum granulosum, with large eosinophilic cytoplasmic inclusions and a perivascular to interstitial lymphohistiocytic infiltration accompanied by fibroblastic proliferation and neovascularization. Negative staining of mucosal tissue homogenates demonstrated parapoxvirus-like particles. The presence of parapoxvirus was confirmed by polymerase chain reaction, using primers specific for parapoxvirus of ungulates. By in situ hybridization, using a parapox-specific, digoxigenin-labeled DNA probe, abundant parapoxvirus DNA-positive epithelial cells were detected in the stratum granulosum and the outer parts of the spiny layer. There was no parapoxvirus-positive signal in the adjacent submucosa. Although DNA analysis revealed that the causative agent can clearly be distinct from terrestrial parapoxviruses, lesions resembled parapoxvirus infections in other terrestrial species, and the pattern of virus DNA distribution indicated a direct effect of the virus on keratinocytes. In contrast, changes in the corium may be considered an indirect response mediated by the virus or the immune system.     

Detection of canine oral papillomavirus-DNA in canine oral squamous cell carcinomas and p53 overexpressing skin papillomas of the dog using the polymerase chain reaction and non-radioactive in situ hybridization

Nineteen cutaneous and mucocutaneous papillomas, as well as 29 oral and 25 non-oral squamous cell carcinomas of dogs were analyzed immunohistologically for the presence of papillomavirus (PV)-antigens. Canine oral papillomavirus (COPV)-DNA was detected in formalin-fixed, paraffin-embedded tissues by polymerase chain reaction (PCR) and non-radioactive in situ hybridization (ISH). Furthermore, the expression of the tumor suppressor protein p53 was investigated. PV-antigens were detectable in more than 50% of the oral and cutaneous papillomas, while no PV-antigens could be demonstrated in venereal papillomas. One squamous cell carcinoma was PV-antigen positive. Only two cutaneous papillomas of the head showed a strong p53-specific immunostaining, while overexpressed p53 was detectable in approximately 35% of all squamous cell carcinomas. It was possible to amplify fragments of the E6, E7 and L1 gene by polymerase chain reaction (PCR) from five of eight oral and from five of eight cutaneous papillomas as well as from three oral squamous cell carcinomas. Nine of 10 papillomas showed a strong nucleus-associated hybridization signal typical for COPV-DNA. In three squamous cell carcinomas COPV-DNA was located in nests of the epithelial tumor cells surrounding ‘horn pearls' or disseminated in the carcinoma tissue. These observations support the view that COPV may also induce non-oral papillomas in the dog and confirm the opinion that a progression of viral papillomas into carcinomas in dogs may occur.     

Digital papillomatosis in a confined Beagle.

Papillomavirus-induced papillomas were diagnosed on multiple digits of all 4 feet of a young Beagle. No other cutaneous or oral involvement was identified. Papillomavirus antigen was confirmed by immunoperoxidase localization within keratinocyte nuclei. In addition to the typical basophilic intranuclear inclusions associated with papillomavirus infections, keratinocytes within the papillomas contained large, eosinophilic cytoplasmic inclusions that previously have been described in a Boxer with cutaneous lesions associated with a papillomavirus infection. The papillomas in this Beagle regressed completely within 2 months of the initial diagnosis.     

Co-occurrence of papillomas related to Equus caballus papillomavirus type 2 and cutaneous habronemiasis

The present report describes a case of cutaneous papillomatosis related to equine papillomavirus type 2 (EcPV2) with concomitant cutaneous habronemiasis. A 2-year-old female Pura Raza Española horse was referred for a dermatological evaluation because of the onset of cutaneous multifocal, nodular lesions on the face characterised by a warty and partially ulcerated overlying epidermis. Multiple biopsies were taken from these lesions, and cutaneous viral papillomas with a concomitant severe focal eosinophilic dermatitis with intralesional Habronematidae nematode larvae were diagnosed. Immunohistochemistry revealed papillomaviral capsid protein L1 in the epidermal proliferative lesion, confirming the presence of a productive infection. Rt-qPCR confirmed the presence of EcPV type 2. To the authors’ knowledge, this is the first report of EcPV2-related papillomas with concurrent habronemiasis.     

Evidence for papillomaviruses in ocular lesions in cattle

Negatively stained preparations of various ocular lesions, generally considered to be precursors to bovine ocular squamous cell carcinoma, were subjected to electron microscopic examination for viruses. Lesions examined included five conjunctival plaques, an acanthotic lesion from an eyelid, 15 conjunctival papillomas, two papillomas of the eyelid and two keratinised elongated proliferative lesions of the eyelid (cutaneous horns). Eight lesions contained particles that resembled papillomaviruses. These consisted of a conjunctival plaque, five conjunctival papillomas, a papilloma of the eyelid and one of the samples of cutaneous horn. The particles were present in small numbers and were found only after prolonged search. The finding of these particles suggests that papillomavirus may be involved in the aetiology of bovine ocular squamous cell carcinoma.     

Detection of papillomavirus DNA in precancerous lesions of the ears of sheep

Sixty-seven benign precancerous cutaneous lesions from the ears of 51 sheep were examined for papillomavirus DNA by hybridisation to radioactively labelled or biotinylated probes of bovine papillomavirus type 1 (BPV 1) DNA under varying conditions of stringency. An additional 16 precancerous lesions from other cutaneous sites on 15 sheep and 15 samples of lesion-free skin from nine sheep were similarly examined. Both total genomic and subgenomic probes were used. DNA from 10 aural lesions and one vulval lesion reacted with the probe in a manner indicative of the presence of episomal papillomavirus DNA. Papillomavirus DNA was detected at low stringency in eight of the 10 aural lesions and the vulval lesions, and at high stringency in two aural lesions. Three out of the 8 aural lesions that were positive at low stringency reacted when re-tested at high stringency. Hybridisation with one of the samples of lesion-free ovine skin produced occasional equivocal signals. One particular positive lesion, an ovine aural cutaneous horn, was studied in more detail. When treated with restriction endonucleases, its restriction enzyme pattern was the same as that for BPV 2 DNA with eight of twelve enzymes and the same as that for BPV 1 DNA with two of the twelve enzymes. It was concluded that this ovine papillomavirus was more closely related to BPV 2 than to BPV 1. The possibility that it could be a subtype of BPV 2 is discussed.     

Papillary squamous cell carcinoma in three young dogs

Papillary squamous cell carcinomas were located on the gingiva of 3 young dogs. The tumors locally invaded the soft tissues of each dog, and invaded bone in 2 dogs. Surgical excision was unsuccessful in eliminating 2 of the tumors. Surgery and radiotherapy were effective, and recurrence has not been observed in 39 months in 1 dog, 32 months in a second, and 10 months in a third. Superficially, the oral masses resembled papillomas, which are known to be caused by viruses. Cytopathologic indication of productive infection was not evident, and papillomavirus antigens could not be detected by immunohistochemical methods. Electron microscopy failed to identify viral particles in 2 of the tumors. High and low molecular weight DNA extracts from 2 of the tumors contained no detectable papillomavirus genome when probed under conditions of either high or low stringency by Southern blot hybridization with a cloned canine oral papillomavirus genome.     

In vitro development and characterization of canine epidermis on a porcine acellular dermal matrix

The aim of this study was to develop and to characterize a canine skin epidermal model able to form a proper epidermis on a porcine acellular dermal matrix (PADM). In addition, the role of fibroblasts in skin barrier formation was studied by incorporating or omitting canine dermal fibroblasts in the PADM. Canine epidermal composites were developed by seeding keratinocytes onto the surface of PADM that were previously seeded or non-seeded with dermal fibroblasts. After 14days of culture under air-exposed conditions and in a special growth medium, skin composites were histologically processed and immunohistochemically characterized to determine the expression of cytokeratins and of vimentin and the presence of basement membrane. In all composites, keratinocytes underwent differentiation to a multilayer epidermis with 5-7 viable cell layers. The stratum basalis, stratum spinosum, stratum granulosum and stratum corneum were identified. The expression of cytokeratins was similar to that described in healthy canine epidermis. Laminin and collagen IV immunostaining revealed a homogeneous layer in the epidermal-dermal junction only when the matrix had been seeded by canine dermal fibroblasts. The model may become a simple, useful and cost-effective tool to investigate the biology and pathology of canine epidermis and could partially replace animal testing in several areas of dermatological research.     

Characterization of canine filaggrin: gene structure and protein expression in dog skin

Background – Filaggrin (FLG) is a key protein for skin barrier formation and hydration of the stratum corneum. In humans, a strong association between FLG gene mutations and atopic dermatitis has been reported. Although similar pathogenesis and clinical manifestation have been argued in canine atopic dermatitis, our understanding of canine FLG is limited. Hypothesis/Objectives – The aim of this study was to determine the structure of the canine FLG gene and to raise anti‐dog FLG antibodies, which will be useful to detect FLG protein in dog skin. Methods – The structure of the canine FLG gene was determined by analysing the publicly available canine genome DNA sequence. Polyclonal anti‐dog FLG antibodies were raised based on the canine FLG sequence analysis and used for defining the FLG expression pattern in dog skin by western blotting and immunohistochemistry. Results – Genomic DNA sequence analysis revealed that canine FLG contained four units of repeated sequences corresponding to FLG monomer protein. Western blots probed with anti‐dog FLG monomer detected two bands at 59 and 54 kDa, which were estimated sizes. The results of immunohistochemistry showed that canine FLG was expressed in the stratum granulosum of the epidermis as a granular staining pattern in the cytoplasmic region. Conclusions and clinical importance – This study revealed the unique gene structure of canine FLG that results in production of FLG monomers larger than those of humans or mice. The anti‐dog FLG antibodies raised in this study identified FLG in dog skin. These antibodies will enable us to screen FLG‐deficient dogs with canine atopic dermatitis or ichthyosis.     

Canine cutaneous epitheliotropic T-cell lymphoma with vesiculobullous lesions resembling human bullous mycosis fungoides

The broad spectrum of clinical signs in canine cutaneous epitheliotropic T-cell lymphoma mimics many inflammatory skin diseases and is a diagnostic challenge. A 13-year-old-male castrated golden retriever crossbred dog presented with multifocal flaccid bullae evolving into deep erosions. A shearing force applied to the skin at the periphery of the erosions caused the epidermis to further slide off the dermis suggesting intraepidermal or subepidermal separation. Systemic signs consisted of profound weight loss and marked respiratory distress. Histologically, the superficial and deep dermis were infiltrated by large, CD3-positive neoplastic lymphocytes and mild epitheliotropism involved the deep epidermis, hair follicle walls and epitrichial sweat glands. There was partial loss of the stratum basale. Bullous lesions consisted of large dermoepidermal and intraepidermal clefts that contained loose accumulations of neutrophils mixed with fewer neoplastic cells in proteinaceous fluid. The lifted epidermis was often devitalized and bordered by hydropic degeneration and partial epidermal collapse. Similar neoplastic lymphocytes formed small masses in the lungs associated with broncho-invasion. Clonal rearrangement analysis of antigen receptor genes in samples from skin and lung lesions using primers specific for canine T-cell receptor gamma (TCRγ) produced a single-sized amplicon of identical sequence, indicating that both lesions resulted from the expansion of the same neoplastic T-cell population. Macroscopic vesiculobullous lesions with devitalization of the lesional epidermis should be included in the broad spectrum of clinical signs presented by canine cutaneous epitheliotropic T-cell lymphoma.