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Abstract: The Papillomaviridae family comprises a large number of viruses that can infect a broad range of hosts including mammals, birds and reptiles giving rise to benign lesions of the skin or mucosal membranes. They are characterized by great genetic diversity yet adhere to common biological principles. In this review, we first describe the structure and function of the viral proteins encoded by papillomaviruses (PVs), with a particular emphasis on bovine papillomaviruses (BPV). We then discuss the role of BPV types 1 and 2 in the pathogenesis of equine sarcoids and present recent evidence implicating a novel equine papillomavirus (EcPV-2) in the pathogenesis of equine genital cancers. [Copyright &y& Elsevier]

Highlights: [•] Chimeric BPV VLP vaccines were constructed to present HIV-1 MPER epitopes. [•] The induction of systemic and mucosal antibody responses was assessed in mice. [•] The induced IgG and sIgA recognized the MPER epitopes presented by HIV-1. [•] IgG and sIgA induced by L1-MPER are able to neutralize clade B and clade C HIV-1. [Copyright &y& Elsevier]

The 44-amino-acid E5 protein of bovine papillomavirus is a dimeric transmembrane protein that exists in a stable complex with the platelet-derived growth factor (PDGF) β receptor, causing receptor activation and cell transformation. The transmembrane domain of the PDGF β receptor is required for complex formation, but it is not known if the two proteins contact one another directly. Here, we studied a PDGF β receptor mutant containing a leucine-to-isoleucine substitution in its transmembrane domain, which prevents complex formation with the wild-type E5 protein in mouse BaF3 cells and inhibits receptor activation by the E5 protein. We selected E5 mutants containing either a small deletion or multiple substitution mutations that restored binding to the mutant PDGF β receptor, resulting in receptor activation and growth factor independence. These E5 mutants displayed lower activity with PDGF β receptor mutants containing other transmembrane substitutions in the vicinity of the original mutation, and one of them cooperated with a receptor mutant containing a distal mutation in the juxtamembrane domain. These results provide strong genetic evidence that the transmembrane domains of the E5 protein and the PDGF β receptor contact one another directly. They also demonstrate that different mutations in the E5 protein allow it to tolerate the same mutation in the PDGF β receptor transmembrane domain and that a mutation in the E5 protein can allow it to tolerate different mutations in the PDGF β receptor. Thus, the rules governing direct interactions between transmembrane helices are complex and not restricted to local interactions. [ABSTRACT FROM AUTHOR]

Seven clinical cases of cutaneous papillomatosis in yaks were studied in Arunachal Pradesh, India. Sporadic, single or a chain of multiple varying size warts appeared around the eyes or on the body. Predominant site of warts was around eyes. Histopathologically, these cases were diagnosed as fibropapilloma. It was confirmed by the detection of BPV-1 and BPV-2 or their mixed infection by PCR and sequencing. Quantitative SYBR Green real-time PCR detected comparatively lower viral DNA copy number in cutaneous warts (CWs). Cases of CWs and its causative agent as bovine papillomavirus (BPVs) are reported for the first time in yaks. [ABSTRACT FROM AUTHOR]

Highlights: [•] We compared two PCR strategies for BPV detection in skin lesions and fluids of bovines. [•] Both FAP and BPV type-specific primer sets could amplify BPV types DNA in skin lesions. [•] BPV type-specific primers were more sensitive than consensus primers. [•] BPV type-specific primers could detect co-infection of BPV. [•] Consensus primers amplified were more suitable to detect putative new BPV types. [Copyright &y& Elsevier]

Bovine papillomaviruses (BPVs) are a diverse group of double-stranded DNA viruses, of which 12 viral types have been detected and characterized so far. However, there is still a limited understanding of the diversity of BPV. Several putative new BPVs have been detected and some of these have been recently characterized as new viral types. However, only a very limited amount of information is available on the pathology associated with these novel viral types yet this information could be of significant value in improving our understanding of the biology of BPV. The objective of this study was to examine some of the epidemiological features of cutaneous bovine papillomatosis in Brazilian cattle, in particular to establish the relationship between BPV types isolated from beef and dairy cattle herds and the lesions they cause. Seventy-two cutaneous lesions were collected from 60 animals. Histopathological, PCR and sequencing assays were conducted to characterize the lesions and detect the BPV types responsible. Phylogenetic analysis was carried out using the maximum likelihood method. BPV types 1-6 and 8-10 were found, as well as a putative new BPV type that belongs to the Deltapapillomavirus genus. The tumors were all classified as fibropapillomas. This is believed to be the first record of BPV types 3 and 10 associated with fibropapillomas. These results confirm that there is a wide range of BPV types that infect cattle, and that an understanding of this diversity is necessary for improved methods of therapeutic treatment. [ABSTRACT FROM AUTHOR]

Efficacy of autogenous vaccine and auto-hemotherapy were tested in cattle suffering from papillomatosis. Five doses of autogenous vaccines were injected on day 1,3,5,7 and 9 to each animal. For auto-hemotherapy, 10 ml of venous blood was Injected intramuscularly on weekly interval for four weeks. The animals given autogenous vaccine alone took two months to recover, while animals given both autogenous vaccine and auto-hemotherapy recovered within one and a half month. Results of the present study suggest that autogenous vaccine is useful for therapeutic purposes in bovine papillomatosis and auto-hemotherapy can serve as an useful adjunct by enhancing regression rate of papillomas. [ABSTRACT FROM AUTHOR]

Bovine cutaneous fibropapillomas are benign skin tumours formed by proliferation of epidermal keratinocytes and dermal fibroblasts caused by bovine papillomaviruses (BPVs). BPV E5 oncoprotein plays a key role in neoplastic cell transformation by specifically binding to the platelet derived growth factor beta receptor (PDGFβR) causing its phosphorylation and activation of proliferation and survival signal transduction pathways, among these phosphatidyl inositol-3-kinase (PI3K)/Akt and Ras-mitogen-activatedprotein-kinase-Erk (Ras-MAPK-Erk) pathways. The aim of this study was to investigate the expression of PDGFβR, its phosphorylation status and expression of the downstream molecules phospho-Akt (pAkt) and phospho-Erk (pErk), in naturally occurring bovine cutaneous fibropapillomas. By immunohistochemistry on serial sections we showed cytoplasmic co-expression of the PDGFβR and E5 protein in neoplastic tissue. Western blot analysis revealed that PDGFβR was phosphorylated in higher amount in tumour samples compared to normal skin. pAkt, but not pErk, was also overexpressed in tumour samples. These findings may provide new insights into the aetiopathogenic mechanisms underlying naturally occurring bovine fibropapillomas and contribute to understanding the molecular scenario underlying BPV induced tumourigenesis. [ABSTRACT FROM AUTHOR]

Papillomaviruses, belonging to the Papillomaviridae family, are small oncogenic viruses, causing papillomas and fibropapillomas in the mucosal and cutaneous epithelia of several animals. In bovine species, thirteen types (BPV 1-13) were characterized to date. In this study, the occurrence of papillomatosis in four outbreaks in cattle herds, coming from Brazilian states were registered. The papillomatous lesions were found located in the teats, udders, head and neck. Under the transmission electron microscope, by the negative staining technique, it was possible to visualized rounded-format papillomavirus, with icosahedral symmetry, characterized as "full" and "empty" particles, measuring on average 60 nm in diameter, in all the 40 samples observed of skin lesion fragments. The ultrathin sections revealed the presence of groups of viral, intranuclear, rounded particles measuring 35 nm in diameter and tubular particles with a diameter of 35-39 nm. At immunoelectron microscopy technique, positivity obtained was marked by the presence of aggregates of viral particles formed by the antigen-antibody interaction. In the immunocytochemistry technique, the antigen-antibody reaction showed colloidal gold particles evenly distributed over the surface of the virus. These results showed the importance of the transmission electron microscopy techniques in the diagnosis of bovine papillomatosis that can be used in routine procedures to identify viral agent of this important disease. [ABSTRACT FROM AUTHOR]

Abstract: The bovine papillomavirus type 1 (BPV1) E2 protein binds as a dimer to the viral genome to promote its transcription, replication and maintenance in keratinocytes. Although BPV1 E2 dimerizes primarily through its DNA-binding domain, it was shown previously that its transactivation domain (TAD) can also dimerize in vitro through formation of a disulfide bond between cysteine 57 (C57) of adjacent monomers and of an ion pair between arginine 172 (R172) and aspartic acid 175 (D175). The function of this TAD dimerization interface in vivo remains unknown. Here, we report the effects of substituting C57, R172 and D175 by alanine on the transactivation activity of BPV E2 as well as on its ability to support viral DNA replication using a novel luciferase-based assay. Results for this mutational analysis suggest that the TAD dimerization interface is not essential for either process but may contribute to the DNA replication activity of BPV1 E2. [Copyright &y& Elsevier]

Background: Papillomaviruses (PVs) are highly epitheliotropic as they usually establish productive infections within squamous epithelia of the skin, the anogenital tract and the oral cavity. In this study, early (E) and late (L) protein expression of bovine papillomavirus type 2 (BPV-2) in the urothelium of the urinary bladder is described in cows and water buffaloes suffering from naturally occurring papillomavirus-associated urothelial bladder tumors. Methods and Findings: E5 protein, the major oncoprotein of the BPV-2, was detected in all tumors. L1 DNA was amplified by PCR, cloned and sequenced and confirmed to be L1 DNA. The major capsid protein, L1, believed to be only expressed in productive papillomavirus infection was detected by Western blot analysis. Immunohistochemical investigations confirmed the presence of L1 protein both in the cytoplasm and nuclei of cells of the neoplastic urothelium. Finally, the early protein E2, required for viral DNA replication and known to be a pivotal factor for both productive and persistent infection, was detected by Western blot and immunohistochemically. Electron microscopic investigations detected electron dense particles, the shape and size of which are consistent with submicroscopic features of viral particles, in nuclei of neoplastic urothelium. Conclusion: This study shows that both active and productive infections by BPV-2 in the urothelium of the bovine and bubaline urinary bladder can occur in vivo. [ABSTRACT FROM AUTHOR]

The article discusses research being done on bovine papular stomatitis virus (BPSV) infections Korean native cattle. It references several studies including "Zoonotic Transmission of Bovine Popular Stomatitis Virus" by P. Holmes, M. Richey, C. Mclnnes, A. Wood, A. Schock, E. Wilson and P. Verney in a 2011 issue of "Veterinary Record" and "Severe Dermatitis on Cattle Teats Caused by Bovine Papular Stomatitis Virus" by Y. Inoshima, T. Nakane and H. Sentsui in a 2009 issue of "Veterinary Record."

Equine sarcoid is the most common tumour of horses and accounts for over half of all equine skin tumours. Six types of sarcoid based on gross appearance and clinical behaviour have been described including occult, verrucous, nodular, fibroblastic, mixed and malevolent. Common locations for sarcoid development include the periocular region, ear pinnae, lips, neck, extremities and ventrum (including groin region). Bovine papillomavirus (BPV) is causally associated with equine sarcoid with genetic haplotype, fly vectors and skin trauma identified as potential risk factors for development of the disease. Histopathology is required for definitive diagnosis of equine sarcoid but incomplete excision is thought to activate latent BPV and stimulate growth. Although there are no uniformly effective treatment options, several modalities have been successful in eliminating or managing equine sarcoid. Surgical excision, intratumoural chemotherapy, cryotherapy, hyperthermia, radiotherapy, immunotherapy and immune modulators are used with degrees of success relative to the accessibility and invasiveness of the tumour. Prevention of equine sarcoid may be facilitated by future development of vaccines against bovine papillomavirus. [ABSTRACT FROM AUTHOR]

Bovine papillomaviruses (BPVs) can infect epithelial cells and fibroblasts, inducing fibropapillomas in cattle. Gap junctions are communication channels between cells composed of connexins (Cxs). This study evaluated expression of Cx26 and the major BPV oncoprotein E5 in bovine cutaneous fibropapillomas. BPV DNA was amplified from 20/20 fibropapillomas and 3/3 samples of normal skin. All fibropapillomas (20/20) were positive by immunostaining for E5, whereas the three normal skin samples were negative. Cx26 was expressed faintly in the normal skin epithelium. Positive cytoplasmic and juxtanuclear immu-noreactivity for Cx26 was evident in 18/20 (90%) fibropapillomas. Western blot analysis demonstrated higher expression of Cx26 in 6/6 fibropapillomas compared to normal skin samples. [ABSTRACT FROM AUTHOR]

Abstract: This report describes the complete genomic sequence and taxonomic position of BPV type 13. The BPV13 genome was amplified using the multiply primed rolling-circle amplification technique and long-template PCR employing two specific primers. The two long PCR fragments obtained were cloned and sequenced via primer walking. The complete genomic sequence of the BPV13 contains 7961bp encoding eight proteins, E1, E2, E4, E5, E6, E7, L1, and L2. Similarly to the E5 gene in BPVs 1 and 2, the putative BPV13 E5 ORF encodes a small transforming protein that contains a hydrophobic transmembrane domain. Meanwhile, the retinoblastoma tumor suppressor-binding domain is absent in the putative BPV13 E7 protein. The presence of these two specific molecular features has been recognized as a distinct marker for the development of fibropapilloma in artiodactyl PV-induced lesions. The phylogenetic analysis demonstrated that BPV13 is a new member of the Deltapapillomavirus genus, to be classified as the third representative of the Delta 4 species. The characterization of the genomic sequence of this novel PV will aid in the interpretation of the pathologies described to be related to this virus and provide support for the development of diagnostic tools for epidemiological surveillance of BPV13 in its potential natural hosts. [Copyright &y& Elsevier]

The p300, CBP, and pCAF lysine acetyltransferase (KAT) proteins have been reported to physically interact with bovine (BPV) and human (HPV) papillomavirus E2 proteins. While overexpression of these KAT proteins enhances E2-dependent transcription, the mechanism has not been determined. Using RNA interference (RNAi) to deplete these factors, we demonstrated that E2 transcriptional activity requires physiological levels of p300, CBP, and pCAF. Each protein appears to have a unique function in E2-dependent transcription, since overexpression of one KAT failed to compensate for RNAi knockdown of another KAT. Using an in vitro acetylation assay, we identified highly conserved lysines that are targeted by p300 for acetylation. The conservative changes of lysines at positions 111 and 112 to arginine were of particular interest. The K111R and the K111R/K112R mutants showed reduced transcriptional activity that was not responsive to p300 overexpression, while the K112R mutant retained activity. p300 and CBP were detected at the viral promoter; however, pCAF was not. We propose a model by which E2 transcriptional activity is controlled by p300-mediated acetylation of lysine 111. This model represents a novel mechanism regulating papillomavirus gene expression. [ABSTRACT FROM AUTHOR]

The article presents a study that described the initial detection of a circular genome deletion in a sample infected with naturally bovine papillomavirus (BPV). It discusses the types, characteristics and classifications of BPV. It explains the process of identifying a novel BPV from epithelial papillomas on the tongue of an infected cattle, quantifying the percentage of BPV-12-del in the BPV genomes in the infected tongue sample, and using Southern Blot to confirm the deleted genome.

Bovine papillomaviruses (BPVs) are oncogenic DNA viruses, which mainly induce benign lesions of cutaneous and/ or mucosal epithelia in cattle. Thirteen (BPV 1-13) different viral genotypes have been characterized so far. BPVs are usually species-specific but BPV 1/2 may also infect equids as well as buffaloes and bison and cause tumors in these species. BPV-induced benign lesions usually regress, however occasionally they develop into cancer particularly in the presence of environmental carcinogenic co-factors. The major transforming protein of BPV is E5, a very short hydrophobic, transmembrane protein with many oncogenic activities. E5 contributes to cell transformation through the activation of the cellular β receptor for the platelet-derived growth factor (PDGFβ-r), it also decreases cell surface expression of major histocompatibility complex class I (MHCI) causing viral escape from immunosurveillance, and plays a role in the inhibition of the intracellular communication by means of aberrant connexin expression. E7 is considered as a weak transforming gene, it synergies with E5 in cell transformation during cancer development. E7 expression correlates in vivo with the over-expression of β1-integrin, which plays a role in the regulation of keratinocytes proliferation and differentiation. Additionally, E7 is involved in cell-mediated immune responses leading to tumour rejection, in anoikis process by direct binding to p600, and in invasion process by upregulation of Matrix metalloproteinase1 (MMP-1) expression. Studies on the role of BPV E5 and E7 oncoproteins in naturally occurring tumours are of scientific value, as they may shed new light on the biological role of these two oncogenes in cell transformation. [ABSTRACT FROM AUTHOR]

The diversity of papillomavirus (PV) found in bovine cutaneous warts from Brazilian cattle was evaluated using the PCR technique with the utilization of consensus primers MY09/11 and by PCR using Bovine Papillomavirus (BPV) type-specific primers followed by sequencing. Eleven cutaneous warts from 6 cattle herds were selected. Six warts were positive for the presence of PV. The presence of BPV types 1, 2, 3, 6 and feline sarcoid-associated PV (FeSarPV) in cutaneous wart lesions, as well as the presence of co-infections, was found. To the best of our knowledge, this is the first time that FeSarPV is described co-infecting a cutaneous wart in Brazil. The present study confirms the previous finding of FeSarPV infecting cattle. These results show the necessity of more studies to investigate the diversity of PV in cattle, its diversity and the possibility of co-infection in cattle and other animals. [ABSTRACT FROM AUTHOR]

Abstract: Equine sarcoids are benign fibroblastic skin tumors affecting equids worldwide. Infection with bovine papillomavirus types 1 and 2 has been implicated as a major fact in the disease development; however, the cellular mechanisms underlying fibroblast transformation are still largely unknown. In the present study, a diagnosis of sarcoid was histologically assessed along with eosinophilic dermatitis. The sarcoid lesion expressed the viral oncoproteins E5 and E2, suggesting a causative role of the virus and its replication. Ribosomal DNA of the nematode Habronema muscae was also revealed in the lesion. This is the first report to describe and discuss an association of cutaneous habronemosis with equine sarcoid. [Copyright &y& Elsevier]

The aim of this study was to evaluate the presence of different types of Bovine papillomavirus (BPV) in cattle skin lesions and to identify new viral types in Brazil. A total of 72 skin lesions were analysed from 66 different bovines by PCR using degenerate and specific primers, and subsequent sequencing. Sequencing quality was determined using Staden package with Phred 30. Similarity analysis was performed with BioEdit and BLAST programs to verify the identity with known BPV types. Phylogenetic analysis was carried out using Maximum Likelihood method with TIM3 + G as nucleotide substitution model in PAUP*, and 1000 non-parametric bootstrap replicates. Analyses revealed the presence of ten different types of BPV in the samples, with the exception of BPV7. The presence of co-infections was very high as almost all samples (89%) were co-infected. A putative new BPV11 subtype was also found in lesions from different animals. These results add significant knowledge about the prevalence and diversity of BPV infection in Brazilian cattle, which could be used in future studies aiming at the development of more specific treatment and diagnostic methods. [ABSTRACT FROM AUTHOR]

Abstract: A new electrochemical DNA biosensor for bovine papillomavirus (BPV) detection that was based on screen-printed electrodes was comprehensively studied by electrochemical methods of cyclic voltammetry (CV) and differential pulse voltammetry (DPV). A BPV probe was immobilised on a working electrode (gold) modified with a polymeric film of poly-L-lysine (PLL) and chitosan. The experimental design was carried out to evaluate the influence of polymers, probe concentration (BPV probe) and immobilisation time on the electrochemical reduction of methylene blue (MB). The polymer poly-L-lysine (PLL), a probe concentration of 1μM and an immobilisation time of 60min showed the best result for the BPV probe immobilisation. With the hybridisation of a complementary target sequence (BPV target), the electrochemical signal decreased compared to a BPV probe immobilised on the modified PLL-gold electrode. Viral DNA that was extracted from cattle with papillomatosis also showed a decrease in the MB electrochemical reduction, which suggested that the decreased electrochemical signal corresponded to a bovine papillomavirus infection. The hybridisation specificity experiments further indicated that the biosensor could discriminate the complementary sequence from the non-complementary sequence. Thus, the results showed that the development of analytical devices, such as a biosensor, could assist in the rapid and efficient detection of bovine papillomavirus DNA and help in the prevention and treatment of papillomatosis in cattle. [Copyright &y& Elsevier]

Bovine papillomavirus type 1 (BPV-1) is an economically important virus that induces tumourigenic pathologies in horses and cows. Given that the BPV-1 L1 major coat protein can self-assemble into highly immunogenic higher-order structures, we transiently expressed it in Nicotiana benthamiana as a prelude to producing a candidate vaccine. It was found that plant codon optimization of L1 gave higher levels of expression than its non-optimized counterpart. Following protein extraction, we obtained high yields (183 mg/kg fresh weight leaf tissue) of relatively pure L1, which had self-assembled into virus-like particles (VLPs). We found that these VLPs elicited a highly specific and strong immune response, and therefore they may have utility as a potential vaccine. This is the first report demonstrating the viable production of a candidate BPV vaccine protein in plants. [ABSTRACT FROM AUTHOR]

In animals and man, some forms of cutaneous papillomas are caused by papillomaviruses. Twelve different genotypes of bovine papillomavirus (BPV) have been identified. BPV-1 through BPV-12 are all strictly species-specific. BPV-1 and BPV-2 are associated with fibropapillomas in cattle; these tumors are formed by excessive proliferation of virus-infected dermal fibroblasts and epidermal keratinocytes. In the present study well known diagnostic procedures including histopathological, immunohistochemical and molecular methods were used for the diagnosis of bovine papillomavirus infection.Samples were taken randomly from five cattle with multiple cutaneous tumor formations for the diagnosis of bovine papillomavirus infection occurring commonly in herds. In the present study five cattle were examined for the presence of multiple cutaneous tumors. Cutaneous fibropapillomatosis were confirmed histopathologically and immunohistochemically. BPV-1 and BPV-2 which are etiological agents for the bovine cutaneous fibropapillomatosis were investigated by PCR. BPV serotyping was performed from five cattle all coming from three different farms. in conclusion papillomaviral DNA was detected by the PCR. The amplified long control region (LCR) DNA sequence was identical to that of BPV-1 and negative for the BPV-2 DNA. When considered economical impact of the disease associated with the abundance of the infection significance ofprotective measures and vaccine studies come forward. [ABSTRACT FROM AUTHOR]

Abstract: The E2 open reading frame of bovine papillomavirus (BPV)-1 encodes a 410 amino acid (aa) transcriptional activator, E2-TA, and collinear polypeptides—E2-TR (243 aa) and E8^E2 (196 aa). E8^E2 and E2-TR share the DNA-binding domain of E2-TA, and both have been defined as transcriptional repressors. Although purified E2-TR and E8^E2 proteins specifically bound E2 sites with similar affinities, only the E2-TR stimulated transcription. Here we show that E2-TR trans-activates E2-dependent promoters 5 to 10-fold in cooperation with cellular factors and in a dose-dependent fashion in epithelial cells and fibroblasts of animal or human origin while E2-TA activated >100-fold and the E8^E2 had no effect. However, in contrast to E2-TA, E2-TR activated transcription from a promoter-proximal position. E2-TR also partially inhibited the BPV-1 P89 or heterologous promoters whereas E8^E2 led to complete repression. Thus, the BPV-1 E2-TR modulates viral gene expression in a manner distinct from other E2 proteins. [Copyright &y& Elsevier]

Enzootic bovine haematuria (EBH), which is known to be caused by the interaction of bracken fern toxin (ptaquiloside) and bovine papillomavirus-2 (BPV-2) is a chronic incurable neoplastic ailment of urinary bladder in cattle in certain regions of the world. The present study was aimed to quantify the carcinogen, ptaquiloside (Pta) and flavonoid, quercetin in various ferns growing in certain EBH enzootic areas of two mountainous states of northern India, Himachal Pradesh (HP) and Uttarakhand, where this ailment is a major problem in hill cattle. Of the 25 studied samples of 16 species of ferns, 9 samples collected from both the states were found to contain Pta on HPLC and LCMS analyses. The concentration of Pta present in the fern samples ranged from 0.00 to 1182.84 µg/g on a dry-matter basis. Further, seven samples revealed the potential to contain Pta. Dryopteris cochleata, Hypodematium crenatum, Pseudocyclosorus canus and Pteris cretica were identified to contain Pta for the first time. Quercetin was also analysed by TLC in 32 fern samples and was detected in 17 samples with its concentration ranging from 0.0% to 0.030%. Samples from HP were found to contain a variable range of quercetin in Christella arida, Deparia japonica, Dryopteris cochleata, Dryopteris juxtaposita, H. crenatum, Polystichum squrrosum and Pteridium revolutum. Similarly, Polystichum squarrosum and Pteris cretica from Uttarakhand contained higher concentration of quercetin. The study also revealed that ferns like D. cochleata, H. crenatum, Onychium tenuifrons, Pseudocyclosorus canus, Pteridium revolutum and Pteris cretica contained both Pta and quercetin. The environmental toxin and flavonoid present in these nonbracken fern species are suspected in the causation of EBH along with BPV-2. [ABSTRACT FROM AUTHOR]

Abstract: The Acrodysplasia (Adp) mutation arises from the insertion of a transgene containing a mouse metallothionein-promoted bovine papilloma virus and human growth hormone-releasing factor gene. Although the transgene is not expressed, mice that are hemizygous for the transgene show skull and paw deformities when the progeny receive the transgene paternally. To elucidate the molecular mechanisms underlying the mutant phenotype and the modified transmission pattern of the Adp phenotype, a junctional fragment around the transgene integration site was cloned. The transgene was inserted into the intronic sequences between exon 3 and exon 4 of the Mdga2 gene and the degree of methylation of the transgene and the severity of the phenotype were reciprocally related in that the transgene was highly or under methylated in normal and deformed mice, respectively. Thus, methylation of the transgene appears to regulate phenotypic expression and imprinting of Adp. [Copyright &y& Elsevier]

Summary This study was conducted with the objectives of detecting bovine papillomavirus type 2 (BPV-2) in urine samples and urinary bladder lesions in bovines using polymerase chain reaction (PCR) and real-time PCR-based molecular diagnostic tests, and quantifying BPV-2 in urinary bladder lesions especially in enzootic bovine haematuria (EBH)-affected animals. BPV-2 viral DNA was detected in urine samples (50%) and urinary bladder tissue (68.6%). Cloning and sequencing results showed a close homology with other Indian BPV-2 sequences. Quantitative real-time PCR (SYBR Green assay) showed that the BPV-2 load was low and similar irrespective of inflammatory or neoplastic lesions in the bladder. It was concluded that BPV-2 DNA is frequently present in urine and urinary bladder lesions in cows in an EBH endemic region and virus load was low in urinary bladder lesions. [ABSTRACT FROM AUTHOR]

Bovine papillomavirus type 12 (BPV-12, putative type BAA1) was detected in epithelial papilloma located on the tongue of an infected cow. Then, the whole genome was sequenced, and phylogenetic analysis illustrated that it should be classified as a member of the genus Xipapillomavirus. The viral genome is 7197 base pairs in length and contains five early ORFs (E1, E2, E4, E7 and E8), three late ORFs (L1, L2 and L3), and a long control region that possesses replication regulatory elements. Meanwhile, mRNA of each gene was detected in the papilloma sample. The papilloma was identified as epithelial papilloma by histological and immunohistochemical examination. Based on the genome information and pathological properties, BAA1 was designated as BPV-12 in this study. [ABSTRACT FROM AUTHOR]

Background: Bovine papillomaviruses (BPVs) types 1 and 2 are the only known papillomaviruses able to jump the species. In fact, BPVs 1/2 induce neoplasia in their natural bovine host but infection is also associated to neoplastic skin lesions in equids termed sarcoids. The equine sarcoid is considered to be the most common equine cutaneous tumour worldwide for which no effective therapy is available. Very little is known about the molecular mechanisms underlying tumourigenesis, although genes contributing to sarcoid development have been identified. Several studies associate the development of cancer to the loss of function of a number of oncosuppressor genes. In this study the putative role of O6-methylguanine-DNA methyltrasferase (MGMT) was investigated for sarcoids. The expression of the oncosuppressor protein was assessed in normal and sarcoid cells and tissues. In addition, the DNA methylation profile was analysed to assess the role of epigenetic mechanism in regulation of MGMT expression. Results: A group of 15 equine sarcoids and two primary sarcoid cell lines (fibroblasts) were analyzed for the expression of MGMT protein by immunohistochemistry, immunofluorescence and Western blotting techniques. The sarcoid cell line EqSO4b and the tumour samples showed a reduction or absence of MGMT expression. To investigate the causes of deregulated MGMT expression, ten samples were analyzed for the DNA methylation profile of the CpG island associated to the MGMT promoter. The analysis of 73 CpGs encompassing the region of interest showed in 1 out of 10 (10%) sarcoids a pronouncedly altered methylation profile when compared to the control epidermal sample. Similarily the EqSO4b cell line showed an altered MGMT methylation pattern in comparison to normal fibroblasts. Conclusion: As previously demonstrated for the oncosuppressor gene FHIT, analysis of MGMT expression in sarcoid tissues and a sarcoid-derived fibroblast cell line further suggests that oncosuppressor silencing may be also involved in BPV-induced equine tumours. Abnormal DNA methylation seems to be one of the possible molecular mechanisms involved in the alteration of MGMT expression. Further studies are required to address other basic molecular mechanisms involved in reduced MGMT expression. This study underlines the possible role of DNA methylation in oncosuppressor inactivation in equine sarcoids. [ABSTRACT FROM AUTHOR]

Abstract: The nucleocytoplasmic partitioning of nanoparticles as a result of cell division is highly relevant to the field of nonviral gene delivery. We reviewed the literature on the intracellular distribution of cell organelles (the endosomal vesicles, Golgi apparatus, endoplasmic reticulum and nucleus), foreign macromolecules (dextrans and plasmid DNA) and inorganic nanoparticles (gold, quantum dot and iron oxide) during mitosis. For nonviral gene delivery particles (lipid- or polymer-based), indirect proof of nuclear entry during mitosis is provided. We also describe how retroviruses and latent DNA viruses take advantage of mitosis to transfer their viral genome and segregate their episomes into the host daughter nuclei. Based on this knowledge, we propose strategies to improve nonviral gene delivery in dividing cells with the ultimate goal of designing nonviral gene delivery systems that are as efficient as their viral counterparts but non-immunogenic, non-oncogenic and easy and inexpensive to prepare. [Copyright &y& Elsevier]

The article discusses about probing Bovine papillomavirus (BPV) in equine sarcoids and inflammatory skin diseases of horses through laser microdissection and genetic amplification. The study was performed on 86 skin biopsies from horses with and without sarcoids, where the biopsies are dissected using lasers and evaluated using genetic amplification by polymerase chain reaction and isothermal loop-mediated amplification. Results claimed that BPV is common in normal and inflamed horse skin.

Abstract: In equids, bovine papillomaviruses of type 1 (BPV-1) and less frequently type 2 induce common, locally aggressive skin tumours termed sarcoids. Whereas BPV infection in cattle usually involves the epidermis and is productive in this skin layer, infection in equids is currently thought to be abortive, with virus solely residing as multiple episomes in dermal fibroblasts. Based on recent observations that do not agree with this assumption, we hypothesised that BPV also infects equid epidermis and is active in this skin layer. To test this hypothesis, we conducted a proof-of-principle study on eight distinct sarcoids. Presence of viral DNA was addressed by qualitative and quantitative BPV-1 PCR from microdissected sarcoid epidermis, and by subsequent amplicon sequencing. Viral activity was assessed by screening sarcoid epidermis for BPV-1 protein expression using immunohistochemistry (IHC) or immunofluorescence (IF). Virus-free equine skin served as negative control throughout the assays. BPV-1 DNA was demonstrated in all sarcoid epidermis samples, with viral DNA loads ranging between 2 and 195copies/cell. Identical BPV-1 E5 genes were identified in epidermis and dermis of each of two sarcoids, yet different E5 variants were found in individual lesions. IHC/IF revealed the presence of E5 and E7 protein in sarcoid epidermis, and L1 capsomers in the squamous layer of one lesion. These findings indicate that BPV infection also involves the epidermis, where it may occasionally be productive. [Copyright &y& Elsevier]

The article discusses a research study on the immunohistochemical and molecular distinction of schwannoma from equine sarcoid. Collected were dermal tumors from 10 horses to determine equine sarcoid as the most common cutaneous neoplasm in horses attributed to bovine papillomavirus (BPV) type 1 or 2 while peripheral nerve sheath tumors (PNST) were also observed on horses. Conclusions indicated that equine dermal neoplasms similar to schwannomas tested through sequencing of BPV and polymerase chain reaction (PCR) have been confiremd as equine sarcoids.

An experiment was conducted to study the preliminary therapeutic effects of anti-tumour WCS peptide in bovine papillomavirus (BPV-1&2)-induced hamster dermal fibroma tumour model. Gross and microscopic changes in peptide treatment at early interval fibroma indicated that growth was arrested while in wellgrown tumours, effect of treatment was not much appreciable. Vascular changes, vacuolar degeneration, mononuclear cell infiltration and inflammatory changes were seen in tumour stroma. Ultrastructural features included presence of fibroblasts and fibrocytes, abundance of banded collagen fibres, enlarged pleomorphic nuclei, marked heterochromatin and abundance of rough endoplasmic reticulum. Certain nuclear and cytoplasmic organelle changes were indicative of therapeutic effects. It may be stated that anti-cancer WCS peptide had better therapeutic effects in early cases of experimentally-induced BPV tumours in hamsters. [ABSTRACT FROM AUTHOR]

Abstract: Bovine digital dermatitis (BDD) is a common infectious foot disease whose aetiology is not fully understood. Its origin is thought to be multifactorial, with treponemes being involved. Using PCR-based techniques, BDD samples from 45 affected cows and intact skin from 8 BDD-affected and 33 healthy cows were assessed for the presence of bovine papillomavirus and Treponema DNA. BPV DNA (mainly BPV-1/2) was detected in 22% of lesions and one skin sample from affected animals, and in 15% (BPV-1/-2) and 23% (BPV-3/4/6/9/10) of skin samples from healthy cows. Using quantitative PCR, Treponema DNA was demonstrated in 38/45 BDD lesions, with bacterial DNA loads ranging between 2×103 and 2.78×105 copies/40ng of total DNA. Qualitative PCR confirmed this result and revealed Treponema DNA in 4 additional BDD samples, thus leading to an overall infection rate of 93.3%. Sequence analysis of amplified Treponema DNA revealed T. pedis sp. nov. in 51%, T. medium ssp. bovis in 37.7%, and T. phagedenis ssp. vaccae in 4.4% of lesions. T. brennaborense was not detected in any of the samples. Six BDD samples contained type IV oral Treponema strains, 6 other harboured so far unpublished Treponema sequences. To our knowledge, this is the first report providing information on BPV infection in BDD-affected cattle, and the Treponema DNA load and occurrence of type IV treponemes in BDD samples. Our findings further support an etiologic association of treponemes, particularly T. pedis sp. nov., with BDD disease, yet indicate that BPVs do not directly contribute to BDD development. [Copyright &y& Elsevier]

Genetic selection from libraries expressing proteins with randomized amino acid segments is a powerful approach to identify proteins with novel biological activities. Here, we assessed the utility of deep DNA sequencing to characterize the composition, diversity, size and stability of such randomized libraries. We used 454 pyrosequencing to sequence a retroviral library expressing small proteins with randomized transmembrane domains. Despite the potential for unintended random mutagenesis during its construction, the overall hydrophobic composition and diversity of the proteins encoded by the sequenced library conformed well to its design. In addition, our sequencing results allowed us to calculate a more accurate estimate of the number of different proteins encoded by the library and suggested that the traditional methods for estimating the size of randomized libraries may overestimate their true size. Our results further demonstrated that no significant genetic bottlenecks exist in the methods used to express complex retrovirus libraries in mammalian cells and recover library sequences from these cells. These findings suggest that deep sequencing can be used to determine the quality and content of other libraries with randomized segments and to follow individual sequences during selection. [ABSTRACT FROM AUTHOR]

Summary: The aetiopathogenesis of urinary bladder tumours in cattle involves prolonged ingestion of bracken fern and infection by bovine papillomavirus types 1 or 2 (BPV-1/2). The oncogenic activity of BPV is largely associated with the major oncoprotein E5. Gap junctions are the only communicating junctions found in animal tissues and are composed of proteins known as connexins. Alterations in connexin expression have been associated with oncogenesis. The present study investigated biochemically and immunohistochemically the expression of connexin 43 in samples of normal (n =2), dysplastic (n =3) and neoplastic (n =23) bovine urothelium. The tumours included 10 carcinomas in situ, five papillary urothelial carcinomas and eight invasive urothelial carcinomas. Normal and dysplastic urothelium had membrane expression of connexin 43, but this was reduced in samples of carcinoma in situ. Papillary urothelial carcinomas showed moderate cytoplasmic and membrane labelling, while invasive carcinoma showed loss of connexin 43 expression. [ABSTRACT FROM AUTHOR]

Abstract: Bovine papillomavirus (BPV)-1 and -2 is linked to equine sarcoids, a commonly observed skin tumor in horses that is of considerable veterinary importance. Previous studies using in situ hybridization have detected BPV DNA only in fibroblasts and not in keratinocytes of sarcoids. In contrast, normal equine skin latently infected with BPV shows a dysplastic epithelium without dermal changes, similar to lesions induced by other papillomavirus types infecting the epithelium. The first goal of our study was to describe the epidermal and dermal characteristics of several stages in sarcoid development. Next, we explored whether BPV can infect epidermal cells in the horse using real-time PCR on laser-micro-dissected keratinocytes and fibroblasts. We found that latently infected normal skin samples and a subset of early stage sarcoids show dysplastic, koilocyte-like epithelial changes. BPV DNA was detected in keratinocytes in 40% of the samples with these particular epithelial properties, whereas advanced sarcoids only had BPV DNA in the fibroblasts. These data may indicate a novel and intriguing pathway of BPV infection in the horse composed of a first step of keratinocyte infection, followed by migration of viral material towards the dermis resulting in infection of sub-epidermal fibroblasts and their fully transformed phenotype. Additionally, an example of co-existence of a dermal BPV-1 and an epidermal BPV-2 infection in the same lesion is shown, indicating that horses can harbor infection with more than one BPV type at the same time. [ABSTRACT FROM AUTHOR]

Abstract: Bovine papillomaviruses (BPV) induce benign tumours of the cutaneous or mucosal epithelia in cattle, but are also involved in the development of cancer of the urinary bladder and of the upper gastrointestinal tract. Current BPV genotyping assays employ techniques developed originally for the detection of human papillomaviruses. These methods rely on consensus PCR amplification and subsequent sequencing and are cumbersome and limited in their analytic sensitivity to detect BPV, especially in multiple infections. In this study, a novel multiplex BPV genotyping assay is described to detect sensitively and specifically BPV-1 to -10 as well as BaPV-11. The assay is based on a multiplex PCR using novel broad-spectrum bovine papillomavirus (BSBP) primers followed by multiplex bovine genotyping (MBG) by Luminex xMAP technology. The detection limit of the assay was shown to be between 10 and 100 BPV genomes. In a first application, BPV was detected in 100% of wart preparations with BPV-8 being most prevalent, followed by types 6, 1 and 10. The majority of warts were positive for at least four BPV types. In conclusion, BSBP-PCR/MBG is a powerful high-throughput method suitable for the study of the natural history of BPV and could be useful to veterinarians for the monitoring of the efficacy of future BPV vaccines. [Copyright &y& Elsevier]

Summary: The aetiopathogenesis of urinary bladder tumours in cattle involves prolonged ingestion of bracken fern and infection by bovine papillomavirus types 1 or 2 (BPV-1/2). E5, the major BPV-1/2 oncoprotein, binds to the activated platelet-derived growth factor β receptor (pPDGF-βR), inducing cell transformation in vitro and spontaneously arising urinary bladder tumours. The aim of this study was to assess whether the 85kDa regulatory subunit (p85) of the phosphatidylinositol-3-kinase (PI3K)-AKT pathway and other transforming signals phospho-JUN (pJUN) and phospho-JUN N-terminal kinases (pJNK) may be important in the development of BPV-associated urothelial carcinomas. A physical interaction between the pPDGF-βR and PI3K was shown in four tumours and two samples of normal bladder tissue by co-immunoprecipitation and western blotting. There was greater expression of the PI3K–AKT–cyclin D3 molecular pathway downstream to the activation of pPDGF-βR in neoplastic compared with normal tissue. pJNK and pJUN were overexpressed in samples of tumour compared with normal mucosal tissue. These findings provide new insights into the aetiopathogenic mechanisms underlying naturally occurring bovine urothelial carcinogenesis and contribute to understanding of the role of E5 oncoprotein in naturally occurring tumorigenesis. [ABSTRACT FROM AUTHOR]

A 3-year-old, mixed breed, male horse showed multiple nodules in different areas of the skin. Lesions occurred predominantly on the lips, cheeks, and submandibular and right inguinal regions. The nodules were characterized as mixed, fibroblastic, verrucous and occult types of sarcoid. Histologically there was proliferation of dermal fibroblasts, with or without pseudoepitheliomatous hyperplasia of the epidermis (frequently ulcerated), and formation of small isolated groups of neoplastic fibroblasts in the superficial dermis. Three tissue samples were submitted to DNA extraction and PCR amplification with generic primers for the internal region of the papillomavirus L1 gene. The amplified products from two samples were sequenced and showed 99% identity with the bovine papillomavirus (BPV) BR- UEL-4. This is the first description of BPV BR-UEL-4 infecting a horse and causing sarcoid in this species. BPV BR-UEL-4 is a putative new BPV type recently identified in skin papillomas in a Brazilian cattle herd. [ABSTRACT FROM AUTHOR]

Abstract: BPV-1 and less commonly BPV-2 are associated with the pathogenesis of equine skin tumours termed sarcoids. We recently documented the transcriptional changes that are induced by BPV-1 in equine fibroblasts using microarray analyses. TLR4 expression was found to be significantly down-regulated by BPV-1. In the present study, we show that TLR4 expression is significantly decreased following the exogenous expression of BPV-1 E2 and E7 in primary equine fibroblasts. The results were confirmed by the demonstration of increased TLR4 expression following siRNA suppression of BPV-1 E2 and E7 viral gene expression. These data imply that BPV-1 is able to subvert the innate immune response by downregulation of TLR4. [Copyright &y& Elsevier]

Summary: Four hundred bovine urothelial tumours and tumour-like lesions were classified in accordance with the 2004 World Health Organization (WHO) morphological classification for human urothelial tumours. The spectrum of neoplastic lesions of the urinary bladder of cattle is becoming wider and bovine urothelial tumours share striking morphological features with their human counterparts. A classification system based on the WHO scheme would also be appropriate for the classification of bovine bladder tumours. Bovine urothelial tumours are most often multiple. Four distinct growth patterns of bovine urothelial tumours and tumour-like lesions are recognized: flat, exophytic or papillary, endophytic and invasive. Carcinoma in situ (CIS) is the most common flat urothelial lesion, accounting for approximately 4% of urothelial tumours. CIS is detected adjacent to papillary and invasive tumours in 80–90% of cases. Approximately 3% of papillary lesions are papillomas and approximately 5% are ‘papillary urothelial neoplasms of low malignant potential’ (PUNLMP). Low-grade carcinoma is the most common urothelial tumour of cattle. High-grade carcinomas, and low and high-grade invasive tumours, are less commonly seen. Bovine papillomavirus (BPV) infection and ingestion of bracken fern both play a central role in carcinogenesis of these lesions. [Copyright &y& Elsevier]

Limited information about the distribution of different bovine papillomavirus (BPV) types in Italy is available; therefore, this study aimed to investigate the presence of BPVs in bovine lesions in the Emilia Romagna region. Sixty-four proliferative lesions were collected between december 2011 and december 2014, and subsequently analysed by qualitative PCR with genus- and type-specific primer pairs, as well as rolling circle amplification (RCA). The results demonstrated, for the first time in Italy, the presence of BPV 6, 7, 8, 10 and 12 and also types previously described elsewhere. In addition, the high prevalence of viral co-infections in this sample set provides new information about viral tropism. [ABSTRACT FROM AUTHOR]

Abstract: The multifunctional papillomavirus E2 protein serves important roles in transcriptional activation and genome maintenance and cooperates with the viral E1 helicase for the initiation of viral DNA replication. The bovine papillomavirus genome contains seventeen E2 binding sites, largely concentrated within the long control region, and a single E1 binding site at the origin of viral replication. Using chromatin immunoprecipitation (ChIP) followed by restriction enzyme digestion and PCR, we show that BPV E1 was present only in the region of an active origin of replication and that BPV E2 remained attached to definable segments of the viral genome at specific stages of the cell cycle. [Copyright &y& Elsevier]

The article presents a study which demonstrates new insights on the pathogenesis of equine sarcoids in horses. The study shows that platelet-derived growth factor β receptor (PDGFβ-r) is more phosphorylated in sarcoid tissue than in normal equine skin using Western blot, and interacts with phosphatidylinositol-3-kinase (P13K), under which P13K-AKT signaling pathway is activated. It also indicates that phosphor-JNK (p-JNK) and phosphor-JUN (p-JUN) are overexpressed in sarcoids.