Modeling Potential Autophagy Pathways in COVID-19 and Sarcoidosis.

Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and mainly affects the lungs. Sarcoidosis is an autoinflammatory disease characterized by the diffusion of granulomas in the lungs and other organs. Here, we discuss how the two diseases might involve some common mechanistic cellular pathways around the regulation of autophagy.

The Induced Expression of BPV E4 Gene in Equine Adult Dermal Fibroblast Cells as a Potential Model of Skin Sarcoid-like Neoplasia.

The equine sarcoid is one of the most common neoplasias in the Equidae family. Despite the association of this tumor with the presence of bovine papillomavirus (BPV), the molecular mechanism of this lesion has not been fully understood. The transgenization of equine adult cutaneous fibroblast cells (ACFCs) was accomplished by nucleofection, followed by detection of molecular modifications using high-throughput NGS transcriptome sequencing. The results of the present study confirm that BPV-E4- and BPV-E1^E4-mediated nucleofection strategy significantly affected the transcriptomic alterations, leading to sarcoid-like neoplastic transformation of equine ACFCs. Furthermore, the results of the current investigation might contribute to the creation of in vitro biomedical models suitable for estimating the fates of molecular dedifferentiability and the epigenomic reprogrammability of BPV-E4 and BPV-E4^E1 transgenic equine ACFC-derived sarcoid-like cell nuclei in equine somatic cell-cloned embryos. Additionally, these in vitro models seem to be reliable for thoroughly recognizing molecular mechanisms that underlie not only oncogenic alterations in transcriptomic signatures, but also the etiopathogenesis of epidermal and dermal sarcoid-dependent neoplastic transformations in horses and other equids. For those reasons, the aforementioned transgenic models might be useful for devising clinical treatments in horses afflicted with sarcoid-related neoplasia of cutaneous and subcutaneous tissues.

[Sarcoidosis--granulomatosis: the modern view of the etiology and pathogenesis with clinical cases review].

Diagnostic criteria of sarcoidosis were offered in the 60-ies of XX century, however today the problem of sarcoidosis is difficult for understanding the different specialists and early detection. The development of laboratory diagnostic of viral infections and introduction of polymerase chain reaction (PCR) has greatly improved the level of diagnosis of herpes infections, reveal the previously unknown etiology of many diseases: sarcoidosis (granulomatosis), migraine, multiple sclerosis, cystic prenatal brain damage, convulsions, Hodgkin's disease and others. Sarcoidosis is the set of clinical symptoms (fatigue, shortness of breath, coughing, heaviness in the chest), laboratory, radiological and histopathological data that allow the doctor to diagnose, predict prognosis and treatment policy. Most often, sarcoidosis affects the lungs and thoracic lymph nodes. In the last period for 2011-2013 in 2930 immunologically tested patients the sarcoidosis was confirmed in 146. Primarily these patients were exposed to different diagnosis--COPD, pneumonia, tuberculosis, lung cancer. Among patients with sarcoidosis on the first place in frequency of detection were EBV and HHV-6. We were the first in 2000, described the Epstein-Barr virus as the causative factor of sarcoidosis, and has been hypothesized the immunopathology of sarcoidosis and principles of individual immunotherapy with a resolution of the granulomatous process in 92 % of cases. Subsequently, this association has been and illustrates the relationship to other viruses (HHV-6, HHV-8) demonstrated by other authors.

Bovine papillomavirus type 13 DNA in equine sarcoids.

Equine sarcoids are locally aggressive fibroblastic neoplasms considered to be the most common skin tumors of horses worldwide. Bovine papillomavirus types 1 and 2 have typically been associated with sarcoids in equids. Investigations aiming to identify papillomavirus strains, aside from bovine papillomaviruses 1 and 2, which might be associated with sarcoid lesions, have been lacking. The aim of this article is to report the identification of a third bovine papillomavirus type, bovine papillomavirus 13, associated with equine sarcoids. Six sarcoid lesions were collected from diverse anatomical sites on two horses from southern Brazil. To detect a broad spectrum of papillomavirus strains, eight degenerate primer pairs designed to detect conserved regions on the L1 and E1 genes were tested on the DNA samples. Direct sequencing was then performed on the obtained amplicons, and sequence identities were compared with sequences from all bovine papillomavirus types. The FAP59/FAP64, MY09/MY11, and AR-E1F2/AR-E1R4 sequences generated from the sarcoids were shown to present 99 to 100% identity with bovine papillomavirus 13, a new bovine papillomavirus type previously described in cattle. The results from this study suggest that there is a need to identify bovine papillomavirus type 13 and other papillomavirus strains that might be associated with sarcoids in diverse geographical areas; such investigations might establish the frequency of occurrence of this viral type in these common tumors of equids.

Sarcoids.

Sarcoids are the most common skin tumors seen in horses worldwide. The pathogenesis of sarcoids is multifactorial, including an association with bovine papillomavirus types 1 and 2 and a genetic susceptibility to tumor development. Clinical manifestations vary and include occult, verrucous, nodular, fibroblastic, mixed, and malignant (malevolent) types. The tumor is nonmetastasizing but can become very aggressive locally. Multiple tumors are common. All clinical types can be present in the same horse. No treatment protocol is universally effective. The tumor has a high risk of recurrence. Recurrent and large tumors are associated with poorer prognoses.

The CD4/CD8 ratio in vitreous fluid is of high diagnostic value in sarcoidosis.

PURPOSE: Sarcoidosis is an idiopathic inflammatory disorder involving multiple organs, and ocular manifestation (represented by granulomatous uveitis) is one of the common features. A well-known immunologic feature in sarcoidosis is an increased CD4+ helper T-cell type 1 lymphocyte subset in bronchoalveolar lavage (BAL) fluid. The current study investigated the vitreous lymphocyte subsets of ocular sarcoidosis to elucidate the immunologic features of this disorder in the eye. DESIGN: Case-control study. PARTICIPANTS AND CONTROLS: Fifty-one eyes of 38 patients with ocular sarcoidosis, confirmed by international diagnostic criteria, were enrolled in this study. Twenty-seven eyes of 26 patients with other causes of uveitis were enrolled as nonsarcoid controls. METHODS: Evaluation of diagnostic tests for cell profiles of ocular sarcoidosis. Lymphocytes in the vitreous samples were analyzed by cytology, polymerase chain reaction, and flow cytometry. Peripheral blood was also obtained from each patient and analyzed in comparison with the vitreous samples. MAIN OUTCOME MEASURES: CD4/CD8 ratios of vitreal and peripheral T lymphocytes. RESULTS: CD4/CD8 ratios of the vitreous T lymphocytes were significantly higher in ocular sarcoidosis than in nonsarcoidosis vitreous samples. In the patients with ocular sarcoidosis, the CD4/CD8 ratios of vitreal T lymphocytes were significantly higher than the CD4/CD8 ratios of peripheral T lymphocytes. No significant differences were found between the CD4/CD8 ratios of vitreal and peripheral T lymphocytes in the patients without sarcoidosis. Moreover, the CD4/CD8 ratios of peripheral T lymphocytes in the patients with ocular sarcoidosis were significantly higher than in patients without sarcoidosis. The sensitivity and specificity of the vitreal CD4/CD8 ratio were 100% and 96.3%, respectively, for the diagnosis of ocular sarcoidosis. CONCLUSIONS: Our findings suggest that the CD4/CD8 ratio of vitreous-infiltrating lymphocytes has high diagnostic value in ocular sarcoidosis, comparable to that of the CD4/CD8 ratio in BAL fluid lymphocytosis for pulmonary sarcoidosis. Furthermore, a high CD4/CD8 ratio of peripheral blood T lymphocytes should be one of the laboratory findings for ocular sarcoidosis. Diagnostic vitrectomy using flow cytometric analysis may be a useful adjunct for the diagnosis of ocular sarcoidosis, particularly in complex cases.

Inoculation of young horses with bovine papillomavirus type 1 virions leads to early infection of PBMCs prior to pseudo-sarcoid formation.

Bovine papillomavirus types 1 and 2 (BPV-1 and BPV-2) are known to induce common equine skin tumours, termed sarcoids. Recently, it was demonstrated that vaccination with BPV-1 virus-like particles (VLPs) is safe and highly immunogenic in horses. To establish a BPV-1 challenge model for evaluation of the protective potential of BPV-1 VLPs, four foals were injected intradermally with infectious BPV-1 virions and with viral genome-based and control inocula, and monitored daily for tumour development. Blood was taken before inoculation and at weekly intervals. BPV-1-specific serum antibodies were detected by a pseudo-virion neutralization assay. Total nucleic acids extracted from tumours, intact skin and PBMCs were tested for the presence of BPV-1 DNA and mRNA using PCR and RT-PCR, respectively. Intralesional E5 oncoprotein expression was determined by immunofluorescence. Pseudo-sarcoids developed exclusively at sites inoculated with virions. Tumours became palpable 11-32 days after virion challenge, reached a size of ≤20 mm in diameter and then resolved in ≤6 months. No neutralizing anti-BPV-1 serum antibodies were detectable pre- or post-challenge. BPV-1 DNA was present in lesions but not in intact skin. In PBMCs, viral DNA was already detectable before lesions were first palpable, in concentrations correlating directly with tumour growth kinetics. PBMCs from two of two foals also harboured E5 mRNA. Immunofluorescence revealed the presence of the E5 protein in tumour fibroblasts, but not in the apparently normal epidermis overlying the lesions. Together with previous findings obtained in horses and cows, these data suggest that papillomavirus infection may include a viraemic phase.

Influenza virus vector iNS1 expressing bovine papillomavirus 1 (BPV1) antigens efficiently induces tumour regression in equine sarcoid patients.

Bovine papillomaviruses types 1 and 2 (BPV1, BPV2) commonly induce skin tumours termed sarcoids in horses and other equids. Sarcoids seriously compromise the health and welfare of affected individuals due to their propensity to resist treatment and reoccur in a more severe form. We have developed influenza (Flu) A and B virus vectors that harbour a truncated NS1 gene (iNS) assuring interferon induction and co-express shuffled BPV1 E6 and E7 antigens for sarcoid immunotherapy. In a safety trial involving 12 healthy horses, intradermal administration of iNSA/E6E7equ and iNSB/E6E7equ was well tolerated, with the only transient side effect being mild fever in four horses. Repeated screening of secretions and faeces by RT-PCR and plaque assay revealed no virus shedding, thus also confirming biological safety. In a patient trial involving 29 horses bearing BPV1-induced single or multiple sarcoids, at least one lesion per horse was intratumourally injected and then boosted with iNSA/E6E7equ and/or iNSB/E6E7equ. The treatment induced a systemic antitumour response as reflected by the synchronous regression of injected and non-injected lesions. Irrespective of vaccination schemes, complete tumour regression was achieved in 10/29 horses. In 10/29 horses, regression is still ongoing (May 2021). Intriguingly, scrapings collected from former tumour sites in two patients tested negative by BPV1 PCR. Nine severely affected individuals with a history of unsuccessful therapeutic attempts did not (6/29) or only transiently (3/29) respond to the treatment. INSA/E6E7equ and iNSB/E6E7equ proved safe and effective in significantly reducing the tumour burden even in severe cases.

The same papillomavirus is present in feline sarcoids from North America and New Zealand but not in any non-sarcoid feline samples.

Feline sarcoids are uncommon dermal neoplasms that are associated with papillomavirus (PV) infection. A single PV type, designated feline sarcoid-associated PV (FeSarPV), was detected in 9 feline sarcoids from North America. As FeSarPV has only been detected within feline sarcoids, the epidemiology of the infection remains unknown. The present study used polymerase chain reaction (PCR) to investigate whether this PV is also present within sarcoids from New Zealand cats. Additionally, as PVs are often host-specific, it was hypothesized that FeSarPV may often asymptomatically infect cats but rarely cause disease. To test this hypothesis, specific PCR primers were designed to investigate the presence of FeSarPV DNA within 120 samples from the skin and mouth of cats without sarcoids. Feline sarcoids from both New Zealand and North America contained FeSarPV DNA sequences. However, FeSarPV DNA was not detected within any non-sarcoid feline sample. To the authors' knowledge, this is the first time that FeSarPV has been reported in a country outside North America. As FeSarPV does not asymptomatically infect cats, feline sarcoids are likely due to cross-species infection. Although the reservoir host of FeSarPV is unknown, the host is present and has contact with cats, in both New Zealand and North America.

Intralesional bovine papillomavirus DNA loads reflect severity of equine sarcoid disease.

REASONS FOR PERFORMING STUDY: Sarcoids are nonmetastasising, yet locally aggressive skin tumours that constitute the most frequent neoplasm in equids. Infection by bovine papillomaviruses types 1 and 2 (BPV-1, BPV-2) has been recognised as major causative factor in sarcoid pathogenesis, but a possible correlation of intralesional virus load with disease severity has not been established thus far. HYPOTHESIS: Given the pathogenic role of BPV-1 and BPV-2 in sarcoid disease, we suggest that intralesional viral DNA concentration may reflect the degree of affection. METHODS: Severity of disease was addressed by recording the tumour growth kinetics, lesion number and tumour type for 37 sarcoid-bearing horses and one donkey. Viral load was estimated via quantitative real-time PCR (qPCR) of the E2, E5, L1 and L2 genes from the BPV-1/-2 genome for one randomly selected lesion per horse and correlated with disease severity. RESULTS: Quantitative PCR against E2 identified viral DNA concentrations ranging from 0-556 copies/tumour cell. Of 16 horses affected by quiescent, slowly growing single tumours or multiple mild-type lesions, 15 showed a viral load up to 1.4 copies per cell. In stark contrast, all equids (22/22) bearing rapidly growing and/or multiple aggressive sarcoids had a viral load between 3 and 569 copies per cell. Consistent results were obtained with qPCR against E5, L1 and L2. CONCLUSIONS: While tumours of the same clinical type carried variable virus load, confirming that viral titre does not determine clinical appearance, we identified a highly significant correlation between intralesional viral load and disease severity. POTENTIAL RELEVANCE: The rapid determination of BPV viral load will give a reliable marker for disease severity and may also be considered when establishing a therapeutic strategy.

Partial sequence analysis of the L1 gene of bovine papillomavirus type 1 detected by PCR with MY09/MY11 primers in equine sarcoids in Poland.

BPV-1 is now recognized as a main etiological agent of equine sarcoids. The etiopathogenesis of the equine sarcoids is equivocal and is not yet fully understood. The aim of the present study was to analyse a partial sequence of the L1 gene of BPV associated with equine sarcoids in Polish horses. After clinical diagnosis, 40 skin lesions obtained from 29 horses were collected. The amplicons of a fragment of BPV L1 DNA were detected using PCR with MY09/MY11 primers in 31 specimens. All of them were recognized as BPV-1. Phylogenetic analysis has allowed the amplicons of partial L1 gene to be divided into 3 phylogenetic groups (A, B, C) and one separate isolate (20c). Sequence variants from phylogenetic groups B, C and isolate 20c represented new genetic variants of BPV-1 L1. Sequence variants from groups B and C were submitted to GenBank NCBI.

Lack of detectable equine herpesviruses 1 and 2 in paraffin-embedded specimens of equine sarcoidosis.

BACKGROUND: Equine sarcoidosis is a rare, multisystemic, noncaseating, granulomatous and lymphoplasmacytic disease of unknown etiology. A recent report described a horse with granulomatous skin disease displaying histologic, electron microscopic, and polymerase chain reaction (PCR) findings consistent with equine herpesvirus 2 (EHV-2). OBJECTIVE: To investigate the presence of EHV-2 and equine herpesvirus 1 (EHV-1) in 8 horses with sarcoidosis. ANIMALS: Eight horses with sarcoidosis, reported previously. METHODS: Retrospective study. PCR assays of the tissues were performed to detect DNA associated with EHV-1 and EHV-2. For both herpesviruses the target was their respective glycoprotein B gene. Positive controls consisted of DNA from viral cultures of culturettes from naturally occurring respiratory infections of EHV-1 and EHV-2. RESULTS: The PCR analyses for both equine herpesviruses' DNA were negative in all 8 horses. CONCLUSION: The failure to detect DNA from EHV-1 and EHV-2 in paraffin-embedded skin of these 8 horses does not discount EHV-1 or EHV-2 as causing some cases of ES, but lends support to the presumably multifactorial etiologic nature of the disease.

Genetic evaluation of bovine papillomavirus types detected in equine sarcoids in Poland.

BACKGROUND: Equine sarcoids are the most common neoplasms in horses. Bovine papilloma- virus type 1 (BPV-1) is the main viral type identified in equine sarcoids in Europe. OBJECTIVE: The aim of the present study was to genetically evaluate BPV types based on DNA analyses of the CDS of the L1 gene. The presence of BPV DNA was confirmed by Degenerate Oligonucleotide-Primed Polymerase Chain Reaction (DOP PCR) with FAP59/FAP64 consensus primers. RESULTS: The DNA was detected in 21/40 (52.5%) of clinically diagnosed sarcoids. More than half of 14 isolates (66.7%) shared 100% homology with BPV-1 Deltapapillomavirus 4 isolate 09 asi UK (Acc. No. MF384289) and 99% nucleotide identity with BPV-1 isolate EqSarc1 (Acc. No. JX678969). A comparison with BPV-1 isolate EqSarc1 revealed one silent mutation in C5827T which did not change the aminoacid codon. The remaining 6 isolates (28.6%) shared 100% nucleotide identity with the BPV-1 (Acc. No. X02346) "wild type" isolate, and 1 isolate (4.8%) demonstrated 99% nucleotide identity with BPV-2 (Acc. No. M20219). CONCLUSIONS: Variants of BPV-1 isolate EqSarc1 (Acc. No. JX678969) constitute the most prevalent type of BPV-1 in Polish horses.

Comprehensive analysis of vitreous specimens for uveitis classification: a prospective multicentre observational study.

PURPOSE: To determine the clinical relevance of vitreous biomarkers in patients with uveitis. DESIGN: Multicentre, prospective, observational study. SETTING: Uveitis outpatient clinics of two academic medical centres in Japan. PATIENT POPULATION: This study included 234 eyes of 191 patients with various uveitis aetiologies: definitive sarcoidosis (61 eyes of 46 patients), suspected sarcoidosis (60 eyes of 45 patients), intraocular tumour (34 eyes of 27 patients), viral infection (20 eyes of 18 patients), non-sarcoidosis (16 eyes of 16 patients) and unknown aetiology (43 eyes of 39 patients). OBSERVATION PROCEDURE: Vitreous samples (taken by pars planta vitrectomy) were analysed with flow cytometry, cytology and multiplex PCR analysis. MAIN OUTCOME MEASURES: The primary outcome measures were the diagnostic values of various biomarkers (T cells, B cells and pathogen DNA) in vitreous samples. The secondary outcome was visual acuity after vitrectomy. RESULTS: Sarcoidosis showed higher CD4/CD8 or CD4+ measurements than other aetiologies (p<0.01). In samples with viral infection, pathogen DNA was detected, and CD8+ counts were higher than the other aetiologies (p<0.01). Eyes with tumour had higher CD19+ (p<0.05). Non-sarcoidosis had lower CD4/CD8 than sarcoidosis, higher CD8+ than sarcoidosis and lower CD19+ than tumour (p<0.01). Unknown uveitis had lower CD4/CD8 than sarcoidosis (p<0.01), and higher CD4/CD8 than non-sarcoidosis, viral infection or tumour (p<0.001). Visual acuity improved after vitrectomy (p<0.001). CONCLUSIONS: Uveitis aetiologies had distinct vitreous biomarker profiles, especially of infiltrating lymphocytes. Analyses of CD4/CD8 ratio, T-lymphocyte and B-lymphocyte subset, and pathogen DNA in vitreous samples have good safety profiles and high diagnostic value for uveitis classification. TRIAL REGISTRATION NUMBER: UMIN000004980; Pre-results.

Autoimmune diseases and HIV infection: A cross-sectional study.

To describe the clinical manifestations, treatments, prognosis, and prevalence of autoimmune diseases (ADs) in human immunodeficiency virus (HIV)-infected patients.All HIV-infected patients managed in the Infectious Diseases Department of the Lyon University Hospitals, France, between January 2003 and December 2013 and presenting an AD were retrospectively included.Thirty-six ADs were found among 5186 HIV-infected patients which represents a prevalence of 0.69% including immune thrombocytopenic purpura (n = 15), inflammatory myositis (IM) (n = 4), sarcoidosis (n = 4), Guillain-Barré syndrome (GBS) (n = 4), myasthenia gravis (n = 2), Graves' disease (n = 2), and 1 case of each following conditions: systemic lupus erythematosus, rheumatoid arthritis, autoimmune hepatitis, Hashimoto thyroiditis and autoimmune hemolytic anemia. One patient presented 2 ADs. Thirty patients were known to be HIV-infected when they developed an AD. The AD preceded HIV infection in 2 patients. GBS and HIV infection were diagnosed simultaneously in 3 cases. At AD diagnosis, CD4 T lymphocytes count were higher than 350/mm in 63% of patients, between 200 and 350/mm in 19% and less than 200/mm in 19%. Twenty patients benefited from immunosuppressant treatments, with a good tolerance.ADs during HIV infection are uncommon in this large French cohort. Immune thrombocytopenic purpura, sarcoidosis, IM, and GBS appear to be more frequent than in the general population. Immunosuppressant treatments seem to be effective and well tolerated.

Severe prolonged red blood cell aplasia and thrombocytopenia induced by parvovirus B19 infection in a patient with sarcoidosis.

We describe an acromegalic patient who developed a parvovirus B19 (PVB19) infection concomitantly with sarcoidosis, which was complicated by chronic red blood cell aplasia and severe thrombocytopenia, despite disappearance of the virus from serum and the production of high levels of specific polyclonal antibodies to PVB19. Mitogen stimulation of the patient's peripheral blood mononuclear cells induced oversecretion of interferon-gamma (IFN-gamma). Because hematopoietic suppression by IFN-gamma has been reported, a possible mechanism underlying reticulocytopenia and thrombocytopenia could involve IFN-gamma.

Detection and quantification of human herpesvirus 6 genomes using bronchoalveolar lavage fluid in immunocompromised patients with interstitial pneumonia.

Human herpesviruses have been recognized as a pathogen involved in interstitial pneumonia (IP), especially in immunocompromised patients. So far, little is known about involvement of human herpesvirus 6 (HHV-6) in systemic respiratory tract disease. Currently, routine diagnostic tests for HHV-6 are inefficient for HHV-6 reactivation, therefore, we established a rapid quantification system of HHV-6 using real-time PCR in order to determine the possible role of human HHV-6 reactivation in immunocompromised patients showing IP. Bronchoalveolar lavage fluid (BALF) specimens were obtained from 84 consecutively treated patients with interstitial lung diseases (ILD) including various types of IP. First, we determined the viral burden in BALF and peripheral blood obtained from healthy volunteers. In healthy volunteers, the prevalence of HHV-6 in BALF was higher (4/12, 33.3%) than in peripheral blood (8/53, 15.1%), ranging from 0 to 101.65 HHV-6 genome copies per 1 microg of DNA. Among 84 patients with ILD analyzed, the prevalence of HHV-6 in BALF was 27.4% (23/84), ranging from 0 to 103.87 copies per 1 microg of DNA. Three specimens obtained from patients with collagen vascular disease, 2 from Hodgkin's disease, and 1 with sarcoidosis had high level of HHV-6 viral DNA, while none of the patients with idiopathic IP showed elevation of HHV-6 (more than 102) in BALF. Our results suggest that measurement of HHV-6 genomes in BALF using real-time PCR may be useful in management of the care of respiratory complications in immunocompromised patients.

Screening for bovine papillomavirus in peripheral blood cells of donkeys with and without sarcoids.

Papillomaviral DNA has been identified in peripheral blood cells of both cattle and humans with and without associated disease and it has been suggested that such cells may act as sites of viral latency. In order to investigate the possibility of latent papillomaviral infection in the aetiopathogenesis of the equine sarcoid, peripheral blood derived DNA samples from 20 healthy and 34 sarcoid-affected donkeys were subject to polymerase chain reaction (PCR) using papillomaviral specific primers. Analysis of blood derived DNA samples failed to demonstrate the presence of papillomaviral DNA in any animal. Screening of 37 matched sarcoid derived DNA samples confirmed the presence of BPV in 34 diseased donkeys. This study supports the hypothesis of BPV as an aetiological agent in the equine sarcoid and suggests that latent virus in circulating peripheral blood cells does not play a role in the pathogenesis and epidemiology of the equine sarcoid.

PCR detection of bovine papilloma virus DNA in superficial swabs and scrapings from equine sarcoids.

The purpose of this study was to examine if bovine papilloma virus (BPV) DNA can be detected in superficial swabs or scrapings from equine sarcoids. Samples were obtained from 92 sarcoids and 20 non-sarcoidal control lesions. The polymerase chain reaction technique was used with a first primer set to check whether DNA extraction was successful, and with a second primer set specific for BPV-DNA. DNA isolation was successful in 88% of the swabs and 93% of the scrapings. All control lesions were negative for BPV-DNA.

Expression of a transforming gene (E5) of bovine papillomavirus in sarcoids obtained from horses.

OBJECTIVE: To determine expression of a transforming gene (E5) of bovine papillomavirus in sarcoids, other tumors, and normal skin samples collected from horses with and without sarcoids. SAMPLE POPULATION: 23 sarcoids and 6 samples of normal skin obtained from 16 horses with sarcoids, 2 samples of normal skin and 2 papillomas obtained from horses without sarcoids, and 1 papilloma obtained from a cow. PROCEDURE: Protein was extracted from tissue samples collected from horses and incubated with agarose beads covalently coupled to Staphylococcus aureus protein A and an anti-E5 polyclonal antibody. Following incubation, proteins were eluted from the beads and electrophoresed on a 14% polyacrylamide gel and transferred to a polyvinylidene difluoride membrane. The E5 protein was detected by use of western blot analysis, using a chemiluminescence detection system. RESULTS: All 23 sarcoids had positive results for expression of E5 protein. Quantity of viral protein appeared to vary among sarcoids. All other tissues examined had negative results for E5 protein. Highest expression for E5 protein was observed in biologically aggressive fibroblastic variants of sarcoids, compared with expression in quiescent tumors. CONCLUSIONS AND CLINICAL RELEVANCE: This study documented that activation and expression of the E5 gene is evident in sarcoids obtained from horses. These data support the conclusion that infection with bovine papillomavirus is important in the initiation or progression of sarcoids in horses. Treatment strategies designed to increase immune recognition of virally infected cells are warranted.