References of "Gillet, Laurent"
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See detailGlycoprotein B cleavage is important for murid herpesvirus 4 to infect myeloid cells.
Glauser, Daniel L.; Milho, Ricardo; Frederico, Bruno et al

in Journal of Virology (2013)

Glycoprotein B (gB) is a conserved herpesvirus virion component implicated in membrane fusion. As with many - but not all - herpesviruses, the gB of murid herpesvirus 4 (MuHV-4) is cleaved into disulfide ... [more ▼]

Glycoprotein B (gB) is a conserved herpesvirus virion component implicated in membrane fusion. As with many - but not all - herpesviruses, the gB of murid herpesvirus 4 (MuHV-4) is cleaved into disulfide-linked subunits, apparently by furin. Preventing gB cleavage for some herpesviruses causes minor infection deficits in vitro, but what the cleavage contributes to host colonization has been unclear. To address this we mutated the furin cleavage site (R-R-K-R) of the MuHV-4 gB. Abolishing gB cleavage did not affect its expression levels, glycosylation or antigenic conformation. In vitro, mutant viruses entered fibroblasts and epithelial cells normally, but had a significant entry deficit in myeloid cells such as macrophages and bone marrow-derived dendritic cells. The deficit in myeloid cells was not due to reduced virion binding or endocytosis, suggesting that gB cleavage promotes infection at a post-endocytic entry step, presumably viral membrane fusion. In vivo, viruses lacking gB cleavage showed reduced lytic spread in the lungs. Alveolar epithelial cell infection was normal, but alveolar macrophage infection was significantly reduced. Normal long-term latency in lymphoid tissue was established nonetheless. [less ▲]

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See detailAlternative splicing switches tropism of a gammaherpesvirus
Machiels, Bénédicte ULiege; Stevenson, P.G.; Vanderplasschen, Alain ULiege et al

Conference (2012, November)

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See detailAlternative splicing switches tropism of a gammaherpesvirus
Machiels, Bénédicte ULiege; Gillet, Laurent ULiege

Conference (2012, November)

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See detailMethod for producing antibody using "naked" expression vector expressing type II transmembrane fusion protein
Renauld, Jean Christophe; Lemaire, Muriel; Dumoutier, Laurie et al

Patent (2012)

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See detailFeeding Cyprinus carpio with infectious materials mediates cyprinid herpesvirus 3 entry through infection of pharyngeal periodontal mucosa
Fournier, Guillaume ULiege; Boutier, Maxime ULiege; Victor, Stalin Raj et al

in Veterinary Research (2012), 43(6),

Cyprinid herpesvirus 3 (CyHV-3), also known as Koi herpesvirus, is the etiological agent of a mortal disease in common and koi carp. Recently, we investigated the entry of CyHV-3 in carp using ... [more ▼]

Cyprinid herpesvirus 3 (CyHV-3), also known as Koi herpesvirus, is the etiological agent of a mortal disease in common and koi carp. Recently, we investigated the entry of CyHV-3 in carp using bioluminescence imaging and a CyHV-3 recombinant strain expressing luciferase (LUC). We demonstrated that the skin is the major portal of entry after inoculation of carp by immersion in water containing CyHV-3. While this model of infection mimics some natural conditions in which infection takes place, other epidemiological conditions could favour entry of virus through the digestive tract. Here, we investigated whether ingestion of infectious materials mediates CyHV-3 entry through the digestive tract. Carp were fed with materials contaminated with the CyHV-3 LUC recombinant (oral contamination) or immersed in water containing the virus (contamination by immersion). Bioluminescence imaging analyses performed at different times post-infection led to the following observations: (i) the pharyngeal periodontal mucosa is the major portal of entry after oral contamination, while the skin is the major portal of entry after contamination by immersion. (ii) Both modes of inoculation led to the spreading of the infection to the various organs tested. However, the timing and the sequence in which some of the organs turned positive were different between the two modes of inoculation. Finally, we compared the disease induced by the two inoculation modes. They led to comparable clinical signs and mortality rate. The results of the present study suggest that, based on epidemiological conditions, CyHV-3 can enter carp either by skin or periodontal pharyngeal mucosal infection. [less ▲]

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See detailMyeloid infection links epithelial and B cell tropisms of murid herpesvirus-4.
Frederico, Bruno; Milho, Ricardo; May, Janet S. et al

in PLoS Pathogens (2012), 8(9), 1002935

Gamma-herpesviruses persist in lymphocytes and cause disease by driving their proliferation. Lymphocyte infection is therefore a key pathogenetic event. Murid Herpesvirus-4 (MuHV-4) is a rhadinovirus that ... [more ▼]

Gamma-herpesviruses persist in lymphocytes and cause disease by driving their proliferation. Lymphocyte infection is therefore a key pathogenetic event. Murid Herpesvirus-4 (MuHV-4) is a rhadinovirus that like the related Kaposi's Sarcoma-associated Herpesvirus persists in B cells in vivo yet infects them poorly in vitro. Here we used MuHV-4 to understand how virion tropism sets the path to lymphocyte colonization. Virions that were highly infectious in vivo showed a severe post-binding block to B cell infection. Host entry was accordingly an epithelial infection and B cell infection a secondary event. Macrophage infection by cell-free virions was also poor, but improved markedly when virion binding improved or when macrophages were co-cultured with infected fibroblasts. Under the same conditions B cell infection remained poor; it improved only when virions came from macrophages. This reflected better cell penetration and correlated with antigenic changes in the virion fusion complex. Macrophages were seen to contact acutely infected epithelial cells, and cre/lox-based virus tagging showed that almost all the virus recovered from lymphoid tissue had passed through lysM(+) and CD11c(+) myeloid cells. Thus MuHV-4 reached B cells in 3 distinct stages: incoming virions infected epithelial cells; infection then passed to myeloid cells; glycoprotein changes then allowed B cell infection. These data identify new complexity in rhadinovirus infection and potentially also new vulnerability to intervention. [less ▲]

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See detailBovine Herpesvirus Type 4 Glycoprotein L Is Nonessential for Infectivity but Triggers Virion Endocytosis during Entry
Lété, Céline ULiege; Machiels, Bénédicte ULiege; Stevenson, P. G. et al

in Journal of Virology (2012)

The core entry machinery of mammalian herpesviruses comprises glycoproteins B, H and L (gB, gH and gL). gH and gL form a heterodimer with a central role in viral membrane fusion. When archetypal alpha- or ... [more ▼]

The core entry machinery of mammalian herpesviruses comprises glycoproteins B, H and L (gB, gH and gL). gH and gL form a heterodimer with a central role in viral membrane fusion. When archetypal alpha- or beta-herpesviruses lack gL, gH misfolds and progeny virions are non-infectious. However, the gL of the rhadinovirus Murid herpesvirus 4 (MuHV-4) is non-essential for infection. In order to define more generally what role gL plays in rhadinovirus infections, we disrupted its coding sequence in Bovine herpesvirus-4 (BoHV-4). BoHV-4 lacking gL showed altered gH glycosylation and incorporated somewhat less gH into virions but remained infectious. However, gL- virions showed poor growth associated with an entry deficit. Moreover a major part of their entry defect appeared to reflect impaired endocytosis, which occurs upstream of membrane fusion itself. Thus, the rhadinovirus gL may be more important for driving virion endocytosis than for incorporating gH into virions, and is non-essential for membrane fusion. [less ▲]

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See detailProteomic characterization of bovine herpesvirus 4 extracellular virions.
Lété, Céline ULiege; Palmeira, Leonor ULiege; Leroy, Baptiste et al

in Journal of Virology (2012), 86(21), 11567-80

Gammaherpesviruses are important pathogens in human and animal populations. During early events of infection, these viruses manipulate preexisting host cell signaling pathways to allow successful ... [more ▼]

Gammaherpesviruses are important pathogens in human and animal populations. During early events of infection, these viruses manipulate preexisting host cell signaling pathways to allow successful infection. The different proteins that compose viral particles are therefore likely to have critical functions not only in viral structures and in entry into target cell but also in evasion of the host's antiviral response. In this study, we analyzed the protein composition of bovine herpesvirus 4 (BoHV-4), a close relative of the human Kaposi's sarcoma-associated herpesvirus. Using mass spectrometry-based approaches, we identified 37 viral proteins associated with extracellular virions, among which 24 were resistant to proteinase K treatment of intact virions. Analysis of proteins associated with purified capsid-tegument preparations allowed us to define protein localization. In parallel, in order to identify some previously undefined open reading frames, we mapped peptides detected in whole virion lysates onto the six frames of the BoHV-4 genome to generate a proteogenomic map of BoHV-4 virions. Furthermore, we detected important glycosylation of three envelope proteins: gB, gH, and gp180. Finally, we identified 38 host proteins associated with BoHV-4 virions; 15 of these proteins were resistant to proteinase K treatment of intact virions. Many of these have important functions in different cellular pathways involved in virus infection. This study extends our knowledge of gammaherpesvirus virions composition and provides new insights for understanding the life cycle of these viruses. [less ▲]

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See detailVirion endocytosis is a major target for Murid Herpesvirus-4 neutralization.
Glauser, D; Gillet, Laurent ULiege; Stevenson, PG

in Journal of General Virology (The) (2012)

Herpesviruses consistently transmit from immunocompetent carriers, implying that their neutralization is hard to achieve. Murid Herpesvirus-4 (MuHV-4) exploits host IgG Fc receptors to bypass blocks to ... [more ▼]

Herpesviruses consistently transmit from immunocompetent carriers, implying that their neutralization is hard to achieve. Murid Herpesvirus-4 (MuHV-4) exploits host IgG Fc receptors to bypass blocks to cell binding, and pH-dependent protein conformation changes to unveil its fusion machinery only after endocytosis. Nevertheless neutralization remains possible by targeting the virion glycoprotein H (gH) / gL heterodimer, and the neutralizing antibody responses of MuHV-4 carriers are improved by boosting with recombinant gH/gL. We analysed here how gH/gL-directed neutralization works. The MuHV-4 gH/gL binds to heparan sulfate. However most gH/gL-specific neutralizing antibodies did not block this interaction. Nor did they act directly on fusion. Instead they blocked virion endocytosis and transport to the late endosomes where membrane fusion normally occurs. The poor endocytosis of gH/gL-neutralized virions was recapitulated precisely by virions genetically lacking gL. Therefore driving virion uptake appears to be an important function of gH/gL that provides a major target for antibody-mediated neutralization. [less ▲]

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See detailGenital re-excretion of Murid gammaherpesvirus 4 following intranasal infection
François, Sylvie ULiege; Vidick, Sarah ULiege; Sarlet, Michaël ULiege et al

in Proceedings of the 1st Scientific Meeting of the Faculty of Veterinary Medicine (2011, December 09)

Gammaherpesviruses are the archetypes of persistent viruses that have been identified in a range of animals from mice to man. As the human gammaviruses have no well-established in vivo infection model ... [more ▼]

Gammaherpesviruses are the archetypes of persistent viruses that have been identified in a range of animals from mice to man. As the human gammaviruses have no well-established in vivo infection model, related animal gammaherpesviruses are an important source of information. We are studying Murid herpesvirus 4 (MuHV-4) in inbred laboratory mouse strains which are commonly accepted as a good model for studying gammaherpesviruses in vivo. To date, it has however never been possible to monitor viral reexcretion and virus transmission in this species. In order to identify potential re-excretion sites, intranasally infected mice were followed through global luciferase imaging for up to six months after infection. Surprisingly, we detected transient viral replication in mice genital tract at various times after latency establishment. Ex vivo imaging, quantitative PCR and immunohistochemistry revealed that virus genomes were present in high quantity in the vaginal tissue and that viral replication occurred mainly at the vaginal external border. Moreover, we highlighted the presence of free infectious viruses in the vaginal cavity at the moment of the observation of viral replication. As this ephemeral viral reexcretion could reveal a link with reproductive cycle, we compared reexcretion in normal and ovariectomized mice. Interestingly, no viral reactivation was observed in absence of hormonal cycle. In conclusion, we experimentally indentified for the first time a reexcretion site for MuHV-4 in mice that had been intranasaly infected. In the future, these results could help us to better understand the biology of gammaherpesviruses but should also allow us to develop strategies that could prevent the spread of these viruses in natural populations. [less ▲]

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See detailGenital re-excretion of Murid gammaherpesvirus 4 following intranasal infection
François, Sylvie ULiege; Vidick, Sarah ULiege; Sarlet, Michaël ULiege et al

Poster (2011, November 16)

Gammaherpesviruses are the archetypes of persistent viruses that have been identified in a range of animals from mice to man. As the human gammaviruses have no well-established in vivo infection model ... [more ▼]

Gammaherpesviruses are the archetypes of persistent viruses that have been identified in a range of animals from mice to man. As the human gammaviruses have no well-established in vivo infection model, related animal gammaherpesviruses are an important source of information. We are studying Murid herpesvirus 4 (MuHV-4) in inbred laboratory mouse strains which are commonly accepted as a good model for studying gammaherpesviruses in vivo. To date, it has however never been possible to monitor viral reexcretion and virus transmission in this species. In order to identify potential re-excretion sites, intranasally infected mice were followed through global luciferase imaging for up to six months after infection. Surprisingly, we detected transient viral replication in mice genital tract at various times after latency establishment. Ex vivo imaging, quantitative PCR and immunohistochemistry revealed that virus genomes were present in high quantity in the vaginal tissue and that viral replication occurred mainly at the vaginal external border. Moreover, we highlighted the presence of free infectious viruses in the vaginal cavity at the moment of the observation of viral replication. As this ephemeral viral reexcretion could reveal a link with reproductive cycle, we compared reexcretion in normal and ovariectomized mice. Interestingly, no viral reactivation was observed in absence of hormonal cycle. In conclusion, we experimentally indentified for the first time a reexcretion site for MuHV-4 in mice that had been intranasaly infected. In the future, these results could help us to better understand the biology of gammaherpesviruses but should also allow us to develop strategies that could prevent the spread of these viruses in natural populations. [less ▲]

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See detailAntibody evasion by a gammaherpesvirus O-Glycan shield
Machiels, Bénédicte ULiege; LETE, Céline ULiege; Guillaume, A. et al

Conference (2011, November)

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See detailAntibody evasion by a gammaherpesvirus O-glycan shield.
Machiels, Bénédicte ULiege; Gillet, Laurent ULiege

Conference (2011, November)

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See detailProfils de liaison et mécanismes d’internalisation des norovirus bovins de génotype 2
Mauroy, Axel ULiege; Gillet, Laurent ULiege; Mathijs, Elisabeth et al

Poster (2011, April)

Appartenant à la famille des Caliciviridae genre Norovirus, les norovirus (NoV) sont des virus non enveloppés dont le génome est composé d’un ARN monocaténaire de polarité positive d’approximativement 7,5 ... [more ▼]

Appartenant à la famille des Caliciviridae genre Norovirus, les norovirus (NoV) sont des virus non enveloppés dont le génome est composé d’un ARN monocaténaire de polarité positive d’approximativement 7,5 kb. Les NoV infectent l’homme et les animaux (bovins, porcins, murins). Chez l’homme, ils sont des agents majeurs de gastroentérite sporadique ou épidémique d’origine souvent alimentaire. Chez le bovin, ils causent également une entérite bénigne et les souches mises en cause sont classées génétiquement dans deux génotypes au sein du génogroupe III du genre Norovirus. La voie d’infection des NoV est oro-fécale, ils sont très résistants dans l’environnement et une infection peut survenir même avec une très faible dose infectieuse. Les NoV humains et animaux sont relativement proches génétiquement et coexistent parfois de manière très étroite en Europe du Nord. Il est donc logique d’envisager le risque zoonotique lié aux NoV animaux et plus particulièrement celui lié aux norovirus bovins (BoNoV). Différents systèmes d’expression protéique ont permis d’obtenir des pseudoparticules virales (VLPs) morphologiquement et antigéniquement semblables à certaines souches de NoV, difficilement cultivables en culture de cellules. Les objectifs du travail étaient d’étudier les types de structures pouvant être impliquées dans la liaison de BoNoVLP aux cellules, ainsi que les mécanismes entrant en jeu pour leur internalisation. Dans une première étude, des VLP d’une souche BoNoV obtenues précédemment ont permis d’investiguer le spectre de liaison des BoNoV de génotype 2 par immunofluorescence indirecte sur des cultures cellulaires bovines d’origines tissulaires différentes. Les VLP ont été capables de se fixer sur chacune des lignées testées tandis que la fluorescence diminuait d’intensité après traitement des cellules par le periodate de sodium. Au cours d’une deuxième étude, les structures permettant l’attachement des BoNoV de génotype 2 ont été investiguées quantitativement en cytométrie en flux avec les VLP. Parmi les enzymes et substances utilisées pour traiter les cellules, le periodate de sodium, l’α-galactosidase, la trypsine, la chymotrypsine et la phospholipase C ont très significativement diminué l’intensité du signal tandis que la neuraminidase a également permis de le réduire modérément. Les sulfates d’héparane ou de chondroïtine n’étaient par contre pas impliqués. Une troisième étude, toujours réalisée en cytométrie en flux, a permis d’évaluer les voies d’internalisation des VLP. Au cours de cette étude, il a été montré que les voies liées aux radeaux lipidiques et de la macropinocytose étaient impliquées. Les trois études menées ont permis de montrer que la structure impliquée dans la liaison des BoNoV de génotype 2 est un saccharide présent sur de nombreux types cellulaires bovins ; que cette structure comprend un résidu α-galactose ; qu’un résidu acide neuraminique peut être aussi impliqué dans cette liaison ou peut la faciliter ; que les sulfates d’héparane et de chondroïtine ne le sont pas ; que des voies alternatives peuvent être utilisées pour l’internalisation de la VLP. Les différents résultats peuvent être intégrés dans le profil de risque zoonotique associé aux BoNoV. [less ▲]

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See detailAntibody evasion by a gammaherpesvirus o-glycan shield.
Machiels, Bénédicte ULiege; Lété, Céline ULiege; Guillaume, Antoine ULiege et al

in PLoS Pathogens (2011), 7(11), 1002387

All gammaherpesviruses encode a major glycoprotein homologous to the Epstein-Barr virus gp350. These glycoproteins are often involved in cell binding, and some provide neutralization targets. However, the ... [more ▼]

All gammaherpesviruses encode a major glycoprotein homologous to the Epstein-Barr virus gp350. These glycoproteins are often involved in cell binding, and some provide neutralization targets. However, the capacity of gammaherpesviruses for long-term transmission from immune hosts implies that in vivo neutralization is incomplete. In this study, we used Bovine Herpesvirus 4 (BoHV-4) to determine how its gp350 homolog - gp180 - contributes to virus replication and neutralization. A lack of gp180 had no impact on the establishment and maintenance of BoHV-4 latency, but markedly sensitized virions to neutralization by immune sera. Antibody had greater access to gB, gH and gL on gp180-deficient virions, including neutralization epitopes. Gp180 appears to be highly O-glycosylated, and removing O-linked glycans from virions also sensitized them to neutralization. It therefore appeared that gp180 provides part of a glycan shield for otherwise vulnerable viral epitopes. Interestingly, this O-glycan shield could be exploited for neutralization by lectins and carbohydrate-specific antibody. The conservation of O-glycosylation sites in all gp350 homologs suggests that this is a general evasion mechanism that may also provide a therapeutic target. [less ▲]

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See detailAlternative attachment factors and internalization pathways for GIII.2 bovine noroviruses.
Mauroy, Axel ULiege; Gillet, Laurent ULiege; Mathijs, Elisabeth ULiege et al

in Journal of General Virology (The) (2011)

Bovine noroviruses belong to the family Caliciviridae, genus Norovirus. Two genotypes are described and viruses genetically related to the Jena and Newbury-2 strains are classified into genotypes 1 and 2 ... [more ▼]

Bovine noroviruses belong to the family Caliciviridae, genus Norovirus. Two genotypes are described and viruses genetically related to the Jena and Newbury-2 strains are classified into genotypes 1 and 2 respectively. In this study, virus-like particles (VLP) of the previously detected B309 Belgian strain, genetically related to genotype 2 bovine noroviruses, were used to investigate virus-host interactions in vitro. B309 VLP were shown to bind to several bovine cell lines. This binding was not affected by heparinase or chondroitinase treatment but was significantly inhibited by both sodium periodate, alpha-galactosidase, trypsin and phospholipase C treatment. Cell treatment by neuraminidase also moderately affected this binding. Taken together, these results show that, in addition to a galactosyl residue, sialic acid could also be involved in binding to susceptible cells. In addition, both the cholesterol-dependent pathway and macropinocytosis are used for B309 VLP internalisation by Madin-Darby Bovine Kidney cells. The data increase the knowledge on bovine norovirus cell interactions. [less ▲]

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See detailEx vivo bioluminescent detection of Alcelaphine herpesvirus 1 infection during malignant catarrhal fever.
Dewals, Benjamin G ULiege; Myster, Françoise ULiege; Palmeira, Leonor ULiege et al

in Journal of Virology (2011), 85(14), 6941-54

Alcelaphine herpesvirus 1 (AlHV-1), carried by wildebeest asymptomatically, causes malignant catarrhal fever (WD-MCF) when cross-species transmitted to a variety of susceptible species of the Artiodactyla ... [more ▼]

Alcelaphine herpesvirus 1 (AlHV-1), carried by wildebeest asymptomatically, causes malignant catarrhal fever (WD-MCF) when cross-species transmitted to a variety of susceptible species of the Artiodactyla order. Experimentally, WD-MCF can be reproduced in rabbits. WD-MCF is described as a combination of lymphoproliferation and degenerative lesions in virtually all organs and caused by unknown mechanisms. Recently, we demonstrated that WD-MCF is associated with the proliferation of CD8(+) cells supporting a latent type of infection in lymphoid tissues. Here, we investigated the macroscopic distribution of AlHV-1 infection using ex vivo bioluminescence imaging in rabbit to determine whether it correlates with the distribution of lesions in lymphoid and non-lymphoid organs. To reach that goal, a recombinant AlHV-1 strain was produced by insertion of a luciferase expression cassette (luc) in an intergenic region. In vitro, the reconstituted AlHV-1 luc(+) strain replicated comparably to the parental strain and luciferase activity was detected by bioluminescence imaging. In vivo, rabbits infected with the AlHV-1 luc(+) strain developed WD-MCF comparably to the parental wild-type strain with hyperthermia and increase of both CD8(+) T cells frequencies and viral genomic charge over time in peripheral blood mononuclear cells and in lymph nodes at time of euthanasia. Bioluminescent imaging revealed that AlHV-1 infection could be detected ex vivo in lymphoid organs but also in lung, liver and kidney during WD-MCF, demonstrating that AlHV-1 infection is prevalent in tissue lesions. Finally, we show that the infiltrating mononuclear leukocytes in non-lymphoid organs are mainly CD8(+) T cells and that latency is predominant during WD-MCF. [less ▲]

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See detailSequencing of Bovine herpesvirus 4 V.test strain reveals important genome features
Palmeira, Leonor ULiege; Machiels, Bénédicte ULiege; Lété, Céline ULiege et al

in Virology Journal (2011), 8(1), 406

Background Bovine herpesvirus 4 (BoHV-4) is a useful model for the human pathogenic gammaherpesviruses Epstein-Barr virus and Kaposi's Sarcoma-associated Herpesvirus. Although genome manipulations of this ... [more ▼]

Background Bovine herpesvirus 4 (BoHV-4) is a useful model for the human pathogenic gammaherpesviruses Epstein-Barr virus and Kaposi's Sarcoma-associated Herpesvirus. Although genome manipulations of this virus have been greatly facilitated by the cloning of the BoHV-4 V.test strain as a Bacterial Artificial Chromosome (BAC), the lack of a complete genome sequence for this strain limits its experimental use. Methods In this study, we have determined the complete sequence of BoHV-4 V.test strain by a pyrosequencing approach. Results The long unique coding region (LUR) consists of 108,241 bp encoding at least 79 open reading frames and is flanked by several polyrepetitive DNA units (prDNA). As previously suggested, we showed that the prDNA unit located at the left prDNA-LUR junction (prDNA-G) differs from the other prDNA units (prDNA-inner). Namely, the prDNA-G unit lacks the conserved pac-2 cleavage and packaging signal in its right terminal region. Based on the mechanisms of cleavage and packaging of herpesvirus genomes, this feature implies that only genomes bearing left and right end prDNA units are encapsulated into virions. Conclusions In this study, we have determined the complete genome sequence of the BAC-cloned BoHV-4 V.test strain and identified genome organization features that could be important in other herpesviruses. [less ▲]

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