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S. A. Holloway - One of the best experts on this subject based on the ideXlab platform.
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genomic variability of Equine Herpesvirus 5
Archives of Virology, 2000Co-Authors: M Dunowska, S. A. Holloway, C R Wilks, J MeersAbstract:Seventeen New Zealand isolates of Equine Herpesvirus 5 (EHV-5) were compared to the Australian prototype strain. PCR primers were designed to amplify EHV-5 glycoprotein B (gB) gene, and Restriction Fragment Length Polymorphism (RFLP) was used to detect differences between cloned PCR products. EHV-5 isolates from different horses showed a high degree of heterogeneity. However, EHV-5 isolates from individual horses remained homogeneous when examined over a period of time or isolated from different sites. A single EHV-5 gB RFLP profile was detected in isolates from each individual horse but one. Two or possibly three different genotypes of EHV-5 were detected in cultures inoculated with a nasal swab of this horse. The heterogeneity observed between EHV-5 isolates from different horses suggests that the use of RFLP may provide a useful epidemiological approach to gain more knowledge about the biology of EHV-5.
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analysis of Equine Herpesvirus 2 strain variation using monoclonal antibodies to glycoprotein b
Archives of Virology, 2000Co-Authors: S. A. Holloway, G. J. Lindquester, M. J. Studdert, H. E. DrummerAbstract:The antigenic relationships of four genomically divergent strains of Equine Herpesvirus 2 (EHV2.86/67, EHV2.5FN, EHV2.141 and EHV2.T-2) and Equine Herpesvirus 5 (EHV5) were examined in ELISA using a panel of EHV2.86/67 gB-specific MAbs. EHV2.86/67 and EHV2.5FN were shown to be more similar to each other than to EHV2.T-2, EHV2.141 or EHV5. Seven of nine EHV2.86/67 gB specific MAbs tested in serum neutralisation assays were shown to neutralise EHV2.86/67 and EHV2.5FN but not EHV2.141, EHV2.T-2 or EHV5. The complete nucleotide and deduced amino acid sequences of EHV2.86/67, EHV2.5FN, EHV2.141 and EHV2.T-2 gB were compared and contrasted with each other and with EHV5 gB. The four EHV2 strains were 94–96% similar at the amino acid level and variability in amino acid sequence mapped to three mains sites designated I, II and III. By contrast, the four EHV2 strains were 77–79% similar to EHV5 gB at the amino acid level. The epitope of these seven gB specific neutralising MAbs has been previously mapped to amino acids 29–74 of EHV2 gB and examination of the deduced amino acid sequence of the four sequenced strains localised the epitope of the seven MAbs to amino acids 30 to 49 located within Site I. Six other divergent strains of EHV2 were examined for variability at Site I using DNA sequencing. Examination of the deduced amino acid sequences of all ten EHV2 strains tested indicated, that based on the epitope of the neutralising MAbs the EHV2 strains formed two distinct antigenic groups, EHV2.86/67-like and EHV2.141-like. EHV5 gB showed divergence from all of the EHV2 gB sequences between amino acids 29–74.
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Identification, sequence analysis and characterisation of Equine Herpesvirus 5 glycoprotein B
Archives of Virology, 1999Co-Authors: S. A. Holloway, G. J. Lindquester, M. J. Studdert, H. E. DrummerAbstract:The complete nucleotide sequence of the gammaHerpesvirus Equine Herpesvirus 5 (EHV5) glycoprotein B (gB) was determined and the deduced amino acid sequence compared with that of the second Equine gammaHerpesvirus EHV2. EHV5 gB is an 870 amino acid protein and is 79% similar and 66% identical with EHV2 gB at the amino acid level. EHV5 gB like EHV2 gB is a disulphide linked heterodimer with subunits of 92 and 68 kDa. EHV5 gB is an integral membrane glycoprotein containing only N-linked oligosaccharides and contains a putative endoproteolytic cleavages site at amino acids 422–485. The EHV5 gB amino acid sequence showed greatest homology with other members of the Rhadinovirus genus of the subfamily Gammaherpesvirinae . Alignment of EHV5 gB sequence with the gB sequence of seven other gammaHerpesviruses showed conservation of 10 cysteine residues as well as conservation of three predicted sites of N-linked glycosylation; the highest degree of conservation of the predicted sites of N-linked glycosylation was observed between EHV5 and the other members of the Rhadinovirus genus. Phylogenetic analysis confirmed EHV2 and EHV5 were most closely related to each other and equally distant from other members of the Rhadinovirus genus included in the analysis.
M. J. Studdert - One of the best experts on this subject based on the ideXlab platform.
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analysis of Equine Herpesvirus 2 strain variation using monoclonal antibodies to glycoprotein b
Archives of Virology, 2000Co-Authors: S. A. Holloway, G. J. Lindquester, M. J. Studdert, H. E. DrummerAbstract:The antigenic relationships of four genomically divergent strains of Equine Herpesvirus 2 (EHV2.86/67, EHV2.5FN, EHV2.141 and EHV2.T-2) and Equine Herpesvirus 5 (EHV5) were examined in ELISA using a panel of EHV2.86/67 gB-specific MAbs. EHV2.86/67 and EHV2.5FN were shown to be more similar to each other than to EHV2.T-2, EHV2.141 or EHV5. Seven of nine EHV2.86/67 gB specific MAbs tested in serum neutralisation assays were shown to neutralise EHV2.86/67 and EHV2.5FN but not EHV2.141, EHV2.T-2 or EHV5. The complete nucleotide and deduced amino acid sequences of EHV2.86/67, EHV2.5FN, EHV2.141 and EHV2.T-2 gB were compared and contrasted with each other and with EHV5 gB. The four EHV2 strains were 94–96% similar at the amino acid level and variability in amino acid sequence mapped to three mains sites designated I, II and III. By contrast, the four EHV2 strains were 77–79% similar to EHV5 gB at the amino acid level. The epitope of these seven gB specific neutralising MAbs has been previously mapped to amino acids 29–74 of EHV2 gB and examination of the deduced amino acid sequence of the four sequenced strains localised the epitope of the seven MAbs to amino acids 30 to 49 located within Site I. Six other divergent strains of EHV2 were examined for variability at Site I using DNA sequencing. Examination of the deduced amino acid sequences of all ten EHV2 strains tested indicated, that based on the epitope of the neutralising MAbs the EHV2 strains formed two distinct antigenic groups, EHV2.86/67-like and EHV2.141-like. EHV5 gB showed divergence from all of the EHV2 gB sequences between amino acids 29–74.
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Identification, sequence analysis and characterisation of Equine Herpesvirus 5 glycoprotein B
Archives of Virology, 1999Co-Authors: S. A. Holloway, G. J. Lindquester, M. J. Studdert, H. E. DrummerAbstract:The complete nucleotide sequence of the gammaHerpesvirus Equine Herpesvirus 5 (EHV5) glycoprotein B (gB) was determined and the deduced amino acid sequence compared with that of the second Equine gammaHerpesvirus EHV2. EHV5 gB is an 870 amino acid protein and is 79% similar and 66% identical with EHV2 gB at the amino acid level. EHV5 gB like EHV2 gB is a disulphide linked heterodimer with subunits of 92 and 68 kDa. EHV5 gB is an integral membrane glycoprotein containing only N-linked oligosaccharides and contains a putative endoproteolytic cleavages site at amino acids 422–485. The EHV5 gB amino acid sequence showed greatest homology with other members of the Rhadinovirus genus of the subfamily Gammaherpesvirinae . Alignment of EHV5 gB sequence with the gB sequence of seven other gammaHerpesviruses showed conservation of 10 cysteine residues as well as conservation of three predicted sites of N-linked glycosylation; the highest degree of conservation of the predicted sites of N-linked glycosylation was observed between EHV5 and the other members of the Rhadinovirus genus. Phylogenetic analysis confirmed EHV2 and EHV5 were most closely related to each other and equally distant from other members of the Rhadinovirus genus included in the analysis.
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Equine Herpesvirus 5 comparisons with ehv2 Equine cytomegalovirus cloning and mapping of a new Equine Herpesvirus with a novel genome structure
Virology, 1992Co-Authors: Catherine Therese Agius, Hadya S Nagesha, M. J. StuddertAbstract:A new Equine Herpesvirus, provisionally designated Equine Herpesvirus 5 (EHV5; Browning and Studdert (1987) J. Gen. Virol. 68, 1441-1447), was examined for the degree of genomic difference from Equine Herpesvirus 2 (EHV2) by Southern hybridizations. EHV5 and EHV2 whole genomic DNA probes were highly specific for homologous DNA only, indicating that significant genomic difference exists between the two viruses. Restriction endonuclease analysis of EHV5 strain 2-141 (EHV5.2-141) revealed that the genome is 179 kb and exists as a single isomer. Clones representing 82% of the genome were obtained and used to construct restriction maps for four restriction endonucleases. Hybridization experiments indicated that the EHV5.2-141 genome does not contain large terminal or internal repeats, although some evidence for very short repeated sequences in the genomic termini was obtained. Such a genome structure makes EHV5 unique among the Equine Herpesviruses but similar to the mouse, rat, and guinea pig cytomegaloviruses and the tupaiid Herpesvirus. Sequence analysis of one of the genomic termini of EHV5.2-141 revealed the presence of a 30-bp sequence (pac-1; Deiss et al. (1986) J. Virol. 59, 605-618) which is highly conserved among Herpesviruses.
H. E. Drummer - One of the best experts on this subject based on the ideXlab platform.
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analysis of Equine Herpesvirus 2 strain variation using monoclonal antibodies to glycoprotein b
Archives of Virology, 2000Co-Authors: S. A. Holloway, G. J. Lindquester, M. J. Studdert, H. E. DrummerAbstract:The antigenic relationships of four genomically divergent strains of Equine Herpesvirus 2 (EHV2.86/67, EHV2.5FN, EHV2.141 and EHV2.T-2) and Equine Herpesvirus 5 (EHV5) were examined in ELISA using a panel of EHV2.86/67 gB-specific MAbs. EHV2.86/67 and EHV2.5FN were shown to be more similar to each other than to EHV2.T-2, EHV2.141 or EHV5. Seven of nine EHV2.86/67 gB specific MAbs tested in serum neutralisation assays were shown to neutralise EHV2.86/67 and EHV2.5FN but not EHV2.141, EHV2.T-2 or EHV5. The complete nucleotide and deduced amino acid sequences of EHV2.86/67, EHV2.5FN, EHV2.141 and EHV2.T-2 gB were compared and contrasted with each other and with EHV5 gB. The four EHV2 strains were 94–96% similar at the amino acid level and variability in amino acid sequence mapped to three mains sites designated I, II and III. By contrast, the four EHV2 strains were 77–79% similar to EHV5 gB at the amino acid level. The epitope of these seven gB specific neutralising MAbs has been previously mapped to amino acids 29–74 of EHV2 gB and examination of the deduced amino acid sequence of the four sequenced strains localised the epitope of the seven MAbs to amino acids 30 to 49 located within Site I. Six other divergent strains of EHV2 were examined for variability at Site I using DNA sequencing. Examination of the deduced amino acid sequences of all ten EHV2 strains tested indicated, that based on the epitope of the neutralising MAbs the EHV2 strains formed two distinct antigenic groups, EHV2.86/67-like and EHV2.141-like. EHV5 gB showed divergence from all of the EHV2 gB sequences between amino acids 29–74.
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Identification, sequence analysis and characterisation of Equine Herpesvirus 5 glycoprotein B
Archives of Virology, 1999Co-Authors: S. A. Holloway, G. J. Lindquester, M. J. Studdert, H. E. DrummerAbstract:The complete nucleotide sequence of the gammaHerpesvirus Equine Herpesvirus 5 (EHV5) glycoprotein B (gB) was determined and the deduced amino acid sequence compared with that of the second Equine gammaHerpesvirus EHV2. EHV5 gB is an 870 amino acid protein and is 79% similar and 66% identical with EHV2 gB at the amino acid level. EHV5 gB like EHV2 gB is a disulphide linked heterodimer with subunits of 92 and 68 kDa. EHV5 gB is an integral membrane glycoprotein containing only N-linked oligosaccharides and contains a putative endoproteolytic cleavages site at amino acids 422–485. The EHV5 gB amino acid sequence showed greatest homology with other members of the Rhadinovirus genus of the subfamily Gammaherpesvirinae . Alignment of EHV5 gB sequence with the gB sequence of seven other gammaHerpesviruses showed conservation of 10 cysteine residues as well as conservation of three predicted sites of N-linked glycosylation; the highest degree of conservation of the predicted sites of N-linked glycosylation was observed between EHV5 and the other members of the Rhadinovirus genus. Phylogenetic analysis confirmed EHV2 and EHV5 were most closely related to each other and equally distant from other members of the Rhadinovirus genus included in the analysis.
Steven R Bolin - One of the best experts on this subject based on the ideXlab platform.
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development and application of a quantitative pcr assay to study Equine Herpesvirus 5 invasion and replication in Equine tissues in vitro and in vivo
Journal of Virological Methods, 2017Co-Authors: Lila M Zarski, Kurt J Williams, Steven R Bolin, Emily A High, Rahul K Nelli, Gisela Soboll HusseyAbstract:Equine Herpesvirus 5 (EHV-5) infection is associated with pulmonary fibrosis in horses, but further studies on EHV-5 persistence in Equine cells are needed to fully understand viral and host contributions to disease pathogenesis. Our aim was to develop a quantitative PCR (qPCR) assay to measure EHV-5 viral copy number in Equine cell cultures, blood lymphocytes, and nasal swabs of horses. Furthermore, we used a recently developed Equine primary respiratory cell culture system to study EHV-5 pathogenesis at the respiratory tract. PCR primers and a probe were designed to target gene E11 of the EHV-5 genome. Sensitivity and repeatability were established, and specificity was verified by testing multiple isolates of EHV-5, as well as DNA from other Equine Herpesviruses. Four-week old fully differentiated (mature), newly seeded (immature) primary Equine respiratory epithelial cell (ERECs), and Equine dermal cell cultures were inoculated with EHV-5 and the cells and supernatants collected daily for 14days. Blood lymphocytes and nasal swabs were collected from horses experimentally infected with Equine Herpesvirus 1 (EHV-1). The qPCR assay detected EHV-5 at stable concentrations throughout 14days in inoculated mature EREC and Equine dermal cell cultures (peaking at 202 and 5861 viral genomes per 106 cellular β actin, respectively). EHV-5 copies detected in the immature EREC cultures increased over 14days and reached levels greater than 10,000 viral genomes per 106 cellular β actin. Moreover, EHV-5 was detected in the lymphocytes of 76% of horses and in the nasal swabs of 84% of horses experimentally infected with EHV-1 pre-inoculation with EHV-1. Post-inoculation with EHV-1, EHV-5 was detected in lymphocytes of 52% of horses while EHV-5 levels in nasal swabs were not significantly different from pre-inoculation levels. In conclusion, qPCR was a reliable technique to investigate viral load in in vivo and in vitro samples, and EHV-5 replication in Equine epithelial cells may be influenced by cellular stages of differentiation.
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Equine multinodular pulmonary fibrosis associated with Equine Herpesvirus 5 in a horse in brazil
Brazilian Journal of Veterinary Pathology, 2014Co-Authors: Welden Panziera, Paula R Giaretta, Glauco J N Galiza, Ailam Lim, Steven R Bolin, Carlos H Borges, Rafael A Fighera, Claudio S L BarrosAbstract:This report describes the first diagnosed case of Equine multinodular pulmonary fibrosis associated with Equine Herpesvirus 5 (EHV-5) in a horse in Brazil. A 7-year-old Thoroughbred mare from a stud farm in southern Brazil died after a chronic respiratory disease characterized by tachypnea, respiratory distress, and cough accompanied by weight loss, anorexia, and intermittent fever. Hematological findings included mild neutrophilia, lymphopenia, and hyperfibrinogenemia. At necropsy the cadaver was extremely emaciated and the lungs did not collapse when the rib cage was removed. Significant gross lesions were restricted to the lungs and consisted of numerous, firm, coalescing nodules widely distributed throughout the organ. The nodules were 1-5 cm in diameter, pale tan-white, and sharply demarcated from the scant normal lung parenchyma. Microscopically, the nodules consisted of marked expansion of the interstitium by wellorganized, mature collagen and infiltration by lymphocytes, macrophages, neutrophils and occasional eosinophils. Cuboidal epithelial cells lined the alveoli in affected areas and their lumina were filled with a moderate to high number of neutrophils and foamy macrophages, which occasionally displayed oval amphophilic intranuclear inclusion bodies. Nucleic acid sequence analyses of amplicons from polymerase chain reaction assays targeting the viral gBand gHgenes showed closest homology with multiple corresponding sequences of the gB and gH genes of EHV 5 available in GenBank.
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experimental induction of pulmonary fibrosis in horses with the gammaHerpesvirus Equine Herpesvirus 5
PLOS ONE, 2013Co-Authors: Kurt J Williams, Ailam Lim, Edward N Robinson, Christina Brandenberger, Roger K Maes, Ashley L Behan, Steven R BolinAbstract:GammaHerpesviruses (γHV) are implicated in the pathogenesis of pulmonary fibrosis in humans and murine models of lung fibrosis, however there is little direct experimental evidence that such viruses induce lung fibrosis in the natural host. The Equine γHV EHV 5 is associated with Equine multinodular pulmonary fibrosis (EMPF), a progressive fibrosing lung disease in its natural host, the horse. Experimental reproduction of EMPF has not been attempted to date. We hypothesized that inoculation of EHV 5 isolated from cases of EMPF into the lungs of clinically normal horses would induce lung fibrosis similar to EMPF. Neutralizing antibody titers were measured in the horses before and after inoculation with EHV 5. PCR and virus isolation was used to detect EHV 5 in antemortem blood and BAL samples, and in tissues collected postmortem. Nodular pulmonary fibrosis and induction of myofibroblasts occurred in EHV 5 inoculated horses. Mean lung collagen in EHV 5 inoculated horses (80 µg/mg) was significantly increased compared to control horses (26 µg/mg) (p < 0.5), as was interstitial collagen (32.6% ± 1.2% vs 23% ± 1.4%) (mean ± SEM; p < 0.001). Virus was difficult to detect in infected horses throughout the experiment, although EHV 5 antigen was detected in the lung by immunohistochemistry. We conclude that the γHV EHV 5 can induce lung fibrosis in the horse, and hypothesize that induction of fibrosis occurs while the virus is latent within the lung. This is the first example of a γHV inducing lung fibrosis in the natural host.
Kelli M Almes - One of the best experts on this subject based on the ideXlab platform.
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identification of Equine Herpesvirus 5 in horses with lymphoma
Journal of Equine Veterinary Science, 2014Co-Authors: Karie Vander Werf, Elizabeth G Davis, Kyathanahalli S Janardhan, Bhupinder Bawa, Steve Bolin, Kelli M AlmesAbstract:Abstract Equine multinodular pulmonary fibrosis, Equine Herpesvirus 5 (EHV-5), and multicentric lymphoma were discovered in one patient. Review of gamma Herpesvirus activity in humans revealed a propensity for lymphoproliferative disorders associated with infection. The objective was to determine the frequency of EHV-5 in lymphoma tissues and compare with the frequency found in the lymph nodes of clinically normal horses. Case control investigation of lymphoma-positive tissues and analysis via polymerase chain reaction (PCR) for EHV-5 was performed on 12 horses. Prospective collection and PCR analysis of lymph nodes (mesenteric or submandibular) for EHV-5 was performed on 21 control horses. Thirteen samples of lymphoma-positive tissues and fluid were submitted for PCR analysis for EHV-5. Of these, 67% was positive. In the control horse population, 14% was positive for EHV-5 ( P = .004). Neoplastic samples positive for EHV-5 were classified as T-cell rich B-cell lymphoma (three), T-cell lymphoma (one), one was nondifferentiated, and two were not stained. Gamma Herpesviruses in humans have been associated with lymphoproliferative diseases such as Kaposi sarcoma and Burkitt lymphoma. This study reveals an increased frequency of EHV-5 (gamma Herpesvirus) in horses diagnosed with lymphoma compared with healthy control horses. Although the exact role this virus plays in the initiation or perpetuation of lymphoproliferative neoplasia is unknown, EHV-5 may be an etiologic agent associated with the development of some types of Equine lymphoma.
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Equine multinodular pulmonary fibrosis and lymphoma in a horse associated with Equine Herpesvirus 5
Journal of Equine Veterinary Science, 2014Co-Authors: Bhupinder Bawa, Karie Vander Werf, Elizabeth G Davis, Laurie A Beard, Gordon A Andrews, Kelli M AlmesAbstract:Abstract A 9-year-old mare presented with a 2-week history of partial anorexia, multiple swellings in the area of the throatlatch, and purulent nasal discharge. On initial presentation, the horse had submandibular and retropharyngeal lymphadenopathy, mild ventral edema, and weight loss. Thoracic radiographs revealed a pulmonary interstitial pattern. Necropsy revealed enlarged lymph nodes throughout the body. The lung parenchyma contained multiple random, well-circumscribed nodules, which, on cut section, were pale, tan colored, and very firm with a distinct line of demarcation from the surrounding normal parenchyma. The subendocardium of the left ventricle, left atrium, and, multifocally, the right ventricle contained white, gritty areas of mineralization. There was marked subintimal mineralization of the aorta and pulmonary artery. Histopathology of the lymph nodes revealed effacement of the parenchyma by a neoplasm composed of large numbers of small mature lymphocytes, fewer large lymphocytes, and scattered moderate numbers of histiocytes. Immunohistochemistry tests for CD3, CD79a, and CD20, confirmed the lymphoma was T-cell-rich B-cell lymphoma. The lungs contained marked interstitial fibrosis with alveolar histiocytosis. Polymerase chain reaction test results of lymph node and lungs were positive for Equine Herpesvirus-5 (EHV-5), a gammaHerpesvirus. GammaHerpesvirus infection has been associated with lymphoma and pulmonary fibrosis in other species. This report describes the association between EHV-5 and both pulmonary fibrosis and lymphoma.