An essential role for gamma-herpesvirus latency-associated nuclear antigen homolog in an acute lymphoproliferative disease of cattle.
; Sorel, Océane ; et al
in Proceedings of the National Academy of Sciences of the United States of America (2013)
Wildebeests carry asymptomatically alcelaphine herpesvirus 1 (AlHV-1), a gamma-herpesvirus inducing malignant catarrhal fever (MCF) to several ruminant species (including cattle). This acute and lethal ... [more ▼]
Wildebeests carry asymptomatically alcelaphine herpesvirus 1 (AlHV-1), a gamma-herpesvirus inducing malignant catarrhal fever (MCF) to several ruminant species (including cattle). This acute and lethal lymphoproliferative disease occurs after a prolonged asymptomatic incubation period after transmission. Our recent findings with the rabbit model indicated that AlHV-1 infection is not productive during MCF. Here, we investigated whether latency establishment could explain this apparent absence of productive infection and sought to determine its role in MCF pathogenesis. First, whole-genome cellular and viral gene expression analyses were performed in lymph nodes of MCF-developing calves. Whereas a severe disruption in cellular genes was observed, only 10% of the entire AlHV-1 genome was expressed, contrasting with the 45% observed during productive infection in vitro. In vivo, the expressed viral genes included the latency-associated nuclear antigen homolog ORF73 but none of the regions known to be essential for productive infection. Next, genomic conformation analyses revealed that AlHV-1 was essentially episomal, further suggesting that MCF might be the consequence of a latent infection rather than abortive lytic infection. This hypothesis was further supported by the high frequencies of infected CD8+ T cells during MCF using immunodetection of ORF73 protein and single-cell RT-PCR approaches. Finally, the role of latency-associated ORF73 was addressed. A lack of ORF73 did not impair initial virus replication in vivo, but it rendered AlHV-1 unable to induce MCF and persist in vivo and conferred protection against a lethal challenge with a WT virus. Together, these findings suggest that a latent infection is essential for MCF induction. [less ▲]Detailed reference viewed: 56 (30 ULg)
A Dose-Effect Relationship For Deltaretrovirus-Dependent Leukemogenesis In Sheep
; ; et al
in Retrovirology (2009), 6Detailed reference viewed: 36 (20 ULg)
Early and transient reverse transcription during primary deltaretroviral infection of sheep.
; ; et al
in Retrovirology (2008), 5
BACKGROUND: Intraindividual genetic variability plays a central role in deltaretrovirus replication and associated leukemogenesis in animals as in humans. To date, the replication of these viruses has ... [more ▼]
BACKGROUND: Intraindividual genetic variability plays a central role in deltaretrovirus replication and associated leukemogenesis in animals as in humans. To date, the replication of these viruses has only been investigated during the chronic phase of the infection when they mainly spread through the clonal expansion of their host cells, vary through a somatic mutation process without evidence for reverse transcriptase (RT)-associated substitution. Primary infection of a new organism necessary involves allogenic cell infection and thus reverse transcription. RESULTS: Here we demonstrate that the primary experimental bovine leukemia virus (BLV) infection of sheep displays an early and intense burst of horizontal replicative dissemination of the virus generating frequent RT-associated substitutions that account for 69% of the in vivo BLV genetic variability during the first 8 months of the infection. During this period, evidence has been found of a cell-to-cell passage of a mutated sequence and of a sequence having undergone both RT-associated and somatic mutations. The detection of RT-dependent proviral substitution was restricted to a narrow window encompassing the first 250 days following seroconversion. CONCLUSION: In contrast to lentiviruses, deltaretroviruses display two time-dependent mechanisms of genetic variation that parallel their two-step nature of replication in vivo. We propose that the early and transient RT-based horizontal replication helps the virus escape the first wave of host immune response whereas somatic-dependent genetic variability during persistent clonal expansion helps infected clones escape the persistent and intense immune pressure that characterizes the chronic phase of deltaretrovirus infection. [less ▲]Detailed reference viewed: 38 (21 ULg)
Vaccination of calves using the BRSV nucleocapsid protein in a DNA prime-protein boost strategy stimulates cell-mediated immunity and protects the lungs against BRSV replication and pathology.
; Boxus, Mathieu ; et al
in Vaccine (2008), 26(37), 4840-8
Respiratory syncytial virus (RSV) is a major cause of respiratory disease in both cattle and young children. Despite the development of vaccines against bovine (B)RSV, incomplete protection and ... [more ▼]
Respiratory syncytial virus (RSV) is a major cause of respiratory disease in both cattle and young children. Despite the development of vaccines against bovine (B)RSV, incomplete protection and exacerbation of subsequent RSV disease have occurred. In order to circumvent these problems, calves were vaccinated with the nucleocapsid protein, known to be a major target of CD8(+) T cells in cattle. This was performed according to a DNA prime-protein boost strategy. The results showed that DNA vaccination primed a specific T-cell-mediated response, as indicated by both a lymphoproliferative response and IFN-gamma production. These responses were enhanced after protein boost. After challenge, mock-vaccinated calves displayed gross pneumonic lesions and viral replication in the lungs. In contrast, calves vaccinated by successive administrations of plasmid DNA and protein exhibited protection against the development of pneumonic lesions and the viral replication in the BAL fluids and the lungs. The protection correlated to the cell-mediated immunity and not to the antibody response. [less ▲]Detailed reference viewed: 53 (11 ULg)
Even attenuated bovine leukemia virus proviruses can be pathogenic in sheep.
Florins, Arnaud-Francois ; Gillet, Nicolas ; Boxus, Mathieu et al
in Journal of virology (2007), 81(18), 10195-200
Based on a reverse genetics approach, we previously reported that bovine leukemia virus (BLV) mutants harboring deletions in the accessory R3 and G4 genes persist at very low proviral loads and are unable ... [more ▼]
Based on a reverse genetics approach, we previously reported that bovine leukemia virus (BLV) mutants harboring deletions in the accessory R3 and G4 genes persist at very low proviral loads and are unable to induce leukemia or lymphoma in sheep, indicating that these R3 and G4 gene sequences are required for pathogenesis. We now show that lymphoma can occur, albeit infrequently (1 case of 20) and after extended periods of latency (7 years). Direct sequencing and reinfection experiments demonstrated that lymphomagenesis was not due to the reversion of the mutant to the wild type. Similar observations with another type of attenuated mutant impaired in the transmembrane protein (TM) YXXL signaling motifs were made. We conclude that the R3 and G4 genes and the TM YXXL motifs are not strictly required for pathogenesis but that their integrity contributes to disease frequency and latency. [less ▲]Detailed reference viewed: 72 (29 ULg)
Cell dynamics and immune response to BLV infection: a unifying model
Florins, Arnaud-Francois ; Gillet, Nicolas ; et al
in Frontiers in Bioscience : A Journal and Virtual Library (2007), 12
Bovine Leukemia virus (BLV) is the natural etiological agent of a lymphoproliferative disease in cattle. BLV can also be transmitted experimentally to a related ruminant species, sheep, in which the ... [more ▼]
Bovine Leukemia virus (BLV) is the natural etiological agent of a lymphoproliferative disease in cattle. BLV can also be transmitted experimentally to a related ruminant species, sheep, in which the pathogenesis is more acute. Although both susceptible species develop a strong anti-viral immune response, the virus persists indefinitely throughout life, apparently at a transcriptionally silent stage, at least in a proportion of infected cells. Soon after infection, these humoral and cytotoxic activities very efficiently abolish the viral replicative cycle, permitting only mitotic expansion of provirus-carrying cells. Short term cultures of these infected cells initially indicated that viral expression protects against spontaneous apoptosis, suggesting that leukemia is a process of accumulation of long-lived cells. This conclusion was recently reconsidered following in vivo dynamic studies based on perfusions of nucleoside (bromodeoxyuridine) or fluorescent protein markers (CFSE). In sheep, the turnover rate of infected cells is increased, suggesting that a permanent clearance process is exerted by the immune system. Lymphocyte trafficking from and to the secondary lymphoid organs is a key component in the maintenance of cell homeostasis. The net outcome of the immune selective pressure is that only cells in which the virus is transcriptionally silenced survive and accumulate, ultimately leading to lymphocytosis. Activation of viral and/or cellular expression in this silent reservoir with deacetylase inhibitors causes the collapse of the proviral loads. In other words, modulation of viral expression appears to be curative in lymphocytic sheep, an approach that might also be efficient in patients infected with the related Human T-lymphotropic virus type 1. In summary, a dynamic interplay between BLV and the host immune response modulates a complex equilibrium between (i) viral expression driving (or) favoring proliferation and (ii) viral silencing preventing apoptosis. As conclusion, we propose a hypothetical model unifying all these mechanisms. [less ▲]Detailed reference viewed: 208 (41 ULg)
DNA immunization with plasmids encoding fusion and nucleocapsid proteins of bovine respiratory syncytial virus induces a strong cell-mediated immunity and protects calves against challenge.
Boxus, Mathieu ; ; et al
in Journal of Virology (2007), 81(13), 6879-89
Respiratory syncytial viruses (RSV) are one of the most important respiratory pathogens of humans and cattle, and there is currently no safe and effective vaccine prophylaxis. In this study, we designed ... [more ▼]
Respiratory syncytial viruses (RSV) are one of the most important respiratory pathogens of humans and cattle, and there is currently no safe and effective vaccine prophylaxis. In this study, we designed two codon-optimized plasmids encoding the bovine RSV fusion (F) and nucleocapsid (N) proteins and assessed their immunogenicity in young calves. Two administrations of both plasmids elicited low antibody levels but primed a strong cell-mediated immunity characterized by lymphoproliferative response and gamma interferon production in vitro and in vivo. Interestingly, this strong cellular response drastically reduced viral replication, clinical signs, and pulmonary lesions after a highly virulent challenge. Moreover, calves that were further vaccinated with a killed-virus vaccine developed high levels of neutralizing antibody and were fully protected following challenge. These results indicate that DNA vaccination could be a promising alternative to the classical vaccines against RSV in cattle and could therefore open perspectives for vaccinating young infants. [less ▲]Detailed reference viewed: 9 (1 ULg)
Development of a 1-step enzyme-linked immunosorbent assay for the rapid diagnosis of bovine respiratory syncytial virus in postmortem specimens.
; ; et al
in Journal of Veterinary Diagnostic Investigation (2007), 19(3), 238-43
Bovine respiratory syncytial virus (BRSV) is associated with severe respiratory disease in cattle. BRSV infection frequently leads to the death of young infected animals. The presence of BRSV in ... [more ▼]
Bovine respiratory syncytial virus (BRSV) is associated with severe respiratory disease in cattle. BRSV infection frequently leads to the death of young infected animals. The presence of BRSV in postmortem specimens is routinely detected using indirect immunofluorescence (IIF). However, this technique requires special equipment and considerable expertise. The present paper describes the development of a 1-step ELISA for rapid (1.5 hours) detection of BRSV antigen in organ homogenates. The performance of the new 1-step ELISA was evaluated using bovine postmortem specimens (n = 108) in comparison with 3 other BRSV diagnostic techniques: indirect immunofluorescence, the Clearview respiratory syncytial virus (RSV) test, and real-time reverse transcriptase polymerase chain reaction (RT-PCR). The relative sensitivity, specificity, and the kappa coefficient of 1-step ELISA, the Clearview RSV electroimmunoassay (EIA), and IIF were calculated, using real-time RT-PCR as the reference test. The new 1-step ELISA was the most sensitive and specific of the 3 tests. Thus, the new 1-step ELISA is a reliable test for detecting BRSV antigen in organ homogenates. [less ▲]Detailed reference viewed: 19 (2 ULg)
Peripheral blood B-cell death compensates for excessive proliferation in lymphoid tissues and maintains homeostasis in bovine leukemia virus-infected sheep.
; Gillet, Nicolas ; et al
in Journal of Virology (2006), 80(19), 9710-9719
The size of a lymphocyte population is primarily determined by a dynamic equilibrium between cell proliferation and death. Hence, lymphocyte recirculation between the peripheral blood and lymphoid tissues ... [more ▼]
The size of a lymphocyte population is primarily determined by a dynamic equilibrium between cell proliferation and death. Hence, lymphocyte recirculation between the peripheral blood and lymphoid tissues is a key determinant in the maintenance of cell homeostasis. Insights into these mechanisms can be gathered from large-animal models, where lymphatic cannulation from individual lymph nodes is possible. In this study, we assessed in vivo lymphocyte trafficking in bovine leukemia virus (BLV)-infected sheep. With a carboxyfluorescein diacetate succinimidyl ester labeling technique, we demonstrate that the dynamics of lymphocyte recirculation is unaltered but that accelerated proliferation in the lymphoid tissues is compensated for by increased death in the peripheral blood cell population. Lymphocyte homeostasis is thus maintained by biphasic kinetics in two distinct tissues, emphasizing a very dynamic process during BLV infection. [less ▲]Detailed reference viewed: 72 (12 ULg)
Estimation du coût global d'un plan de lutte contre l'IBR en Belgique
; ; Muylkens, Benoît et al
in Epidémiologie et Santé Animale (2006), 49Detailed reference viewed: 10 (4 ULg)
Valproate activates bovine leukemia virus gene expression, triggers apoptosis, and induces leukemia/lymphoma regression in vivo.
; Florins, Arnaud-Francois ; Gillet, Nicolas et al
in Proceedings of the National Academy of Sciences of the United States of America (2005), 102(29), 10309-14
Leukemogenic viruses like human T-lymphotropic virus and bovine leukemia virus (BLV) presumably persist in the host partly by latent integration of the provirus in a fraction of infected cells, leading to ... [more ▼]
Leukemogenic viruses like human T-lymphotropic virus and bovine leukemia virus (BLV) presumably persist in the host partly by latent integration of the provirus in a fraction of infected cells, leading to accumulative increase in the outgrowth of transformed cells. Furthermore, viral infection also correlates with a blockade of the apoptotic mechanisms concomitant with an apparent latency of the host cell. Conceptually, induction of viral or cellular gene expression could thus also be used as a therapeutic strategy against retroviral-associated leukemia. Here, we provide evidence that valproate, an inhibitor of deacetylases, activates BLV gene expression in transient transfection experiments and in short-term cultures of primary B-lymphocytes. In vivo, valproate injection into newly BLV-inoculated sheep did not abrogate primary infection. However, valproate treatment, in the absence of any other cytotoxic drug, was efficient for leukemia/lymphoma therapy in the sheep model leading to decreased lymphocyte numbers (respectively from 25.6, 35.7, and 46.5 x 10(3) cells per mm3 to 1.0, 10.6, and 24.3 x 10(3) cells per mm3 in three leukemic sheep) and tumor regression (from >700 cm3 to undetectable). The concept of a therapy that targets the expression of viral and cellular genes might be a promising treatment of adult T cell leukemia or tropical spastic paraparesis/human T-lymphotropic virus-associated myelopathy, diseases for which no satisfactory treatment exists so far. [less ▲]Detailed reference viewed: 81 (31 ULg)
Fate of premalignant clones during the asymptomatic phase preceding lymphoid malignancy.
; ; et al
in Cancer Research (2005), 65(4), 1234-43
Almost all cancers are preceded by a prolonged period of clinical latency during which a combination of cellular events helps move carcinogen-exposed cells towards a malignant phenotype. Hitherto ... [more ▼]
Almost all cancers are preceded by a prolonged period of clinical latency during which a combination of cellular events helps move carcinogen-exposed cells towards a malignant phenotype. Hitherto, investigating the fate of premalignant cells in vivo remained strongly hampered by the fact that these cells are usually indistinguishable from their normal counterparts. Here, for the first time, we have designed a strategy able to reconstitute the replicative history of the bona fide premalignant clone in an animal model, the sheep experimentally infected with the lymphotropic bovine leukemia virus. We have shown that premalignant clones are early and clearly distinguished from other virus-exposed cells on the basis of their degree of clonal expansion and genetic instability. Detectable as early as 0.5 month after the beginning of virus exposure, premalignant cells displayed a two-step pattern of extensive clonal expansion together with a mutation load approximately 6 times higher than that of other virus-exposed cells that remained untransformed during the life span of investigated animals. There was no fixation of somatic mutations over time, suggesting that they regularly lead to cellular death, partly contributing to maintain a normal lymphocyte count during the prolonged premalignant stage. This equilibrium was finally broken after a period of 18.5 to 60 months of clinical latency, when a dramatic decrease in the genetic instability of premalignant cells coincided with a rapid increase in lymphocyte count and lymphoma onset. [less ▲]Detailed reference viewed: 13 (9 ULg)
Genetic immunisation of cattle against bovine herpesvirus 1: glycoprotein gD confers higher protection than glycoprotein gC or tegument protein VP8.
; ; et al
in Veterinary Research (2005), 36(4), 529-44
Bovine herpesvirus 1 (BoHV-1) has frequently been used as a model for testing parameters affecting DNA immunisation in large animals like cattle. However, the selection of target antigens has been poorly ... [more ▼]
Bovine herpesvirus 1 (BoHV-1) has frequently been used as a model for testing parameters affecting DNA immunisation in large animals like cattle. However, the selection of target antigens has been poorly studied, and most of the experiments have been conducted in mice. In the present study, we demonstrated in cattle that a DNA vaccine encoding BoHV-1 glycoprotein gD induces higher neutralising antibody titres than vaccines encoding BoHV-1 gC. Additionally, we show that a DNA vaccine encoding a secreted form of gD induces a higher immune response than a vaccine encoding full-length gD. However, the enhanced immunogenicity associated with the secretion of gD could not be extended to the glycoprotein gC. The current study also describes for the first time the development and the evaluation of a DNA vaccine encoding the major tegument protein VP8. This construct, which is the first BoHV-1 plasmid vaccine candidate that is not directed against a surface glycoprotein, induced a high BoHV-1 specific cellular immunity but no humoral immune response. The calves vaccinated with the constructs encoding full-length and truncated gD showed a non-significant tenfold reduction of virus excretion after challenge. Those calves also excreted virus for significantly (p < 0.05) shorter periods (1.5 days) than the non-vaccinated controls. The other constructs encoding gC and VP8 antigens induced no virological protection as compared to controls. Altogether the DNA vaccines induced weaker immunity and protection than conventional marker vaccines tested previously, confirming the difficulty to develop efficient DNA vaccines in large species. [less ▲]Detailed reference viewed: 27 (7 ULg)
Involvement of glutathione as a mechanism of indirect protection against spontaneous ex vivo apoptosis associated with bovine leukemia virus.
; ; et al
in Journal of virology (2004), 78(12),Detailed reference viewed: 19 (7 ULg)
Conditional expression of type I interferon-induced bovine Mx1 GTPase in a stable transgenic vero cell line interferes with replication of vesicular stomatitis virus
Baise, Etienne ; ; et al
in Journal of Interferon & Cytokine Research (2004), 24(9), 513-521
In some vertebrate species, type I interferon(IFN)-induced Mx gene expression has been shown to confer resistance to some single-stranded RNA (ssRNA) viruses in vitro. Because the bovine species is ... [more ▼]
In some vertebrate species, type I interferon(IFN)-induced Mx gene expression has been shown to confer resistance to some single-stranded RNA (ssRNA) viruses in vitro. Because the bovine species is subject to an exceptionally wide array of infections caused by such viruses, it is anticipated that an antiviral allele should have been retained by evolution at the bovine Mx locus. The identification of such allele may help in evaluating the real significance of the Mx genotype for disease resistance in vivo, in deciphering host-virus molecular interactions involved, or in improving innate disease resistance of livestock through marker-assisted selection. We validated a double transgenic Vero cell clone in which the bovine Mx1 reference allele is placed under control of the human cytomegalovirus (CMV) enhancer-promoter sequence containing elements from the bacterial tetracycline resistance operon to regulate transcription. In the selected clone, transgene repression was very tight, and derepression by doxycycline led to homogeneous 48-h duration expression of physiologic levels of bovine Mx1. Expression of the transgene caused a dramatic decrease in cytopathic efficiency and a 500-5000-fold yield reduction of the Indiana and New Jersey serotypes of vesicular stomatitis virus (VSV). To our knowledge, the transgenic clone developed here is the first ever reported that allows conditional expression of an Mx protein, thus providing a valuable tool for studying functions of Mx proteins in general and that of bovine Mx1 in particular. This latter may henceforward be included in the group of Mx proteins with authenticated anti-VSV activity, which offers new research avenues into the field of host-virus interactions. [less ▲]Detailed reference viewed: 43 (14 ULg)
Reduced proviral loads during primo-infection of sheep by Bovine Leukemia virus attenuated mutants.
; ; et al
in Retrovirology (2004), 1(1),Detailed reference viewed: 13 (2 ULg)
Mesure de l'effet de l'hyperimmunisation pour le contrôle de la rhinotrachéite infectieuse bovine dans les troupeaux laitiers en Belgique : analyse de survie
; ; Thiry, Etienne
in Epidémiologie et Santé Animale (2003), 44Detailed reference viewed: 11 (1 ULg)
Increased cell proliferation-but not reduced cell death-induces lymphocytosis in Bovine Leukaemia Virus-infected sheep
; ; et al
in Abstracts of papers presented at the 2002 meeting of retroviruses (2002, May)Detailed reference viewed: 5 (1 ULg)
In vivo cell turnover in BLV-infected sheep.
; ; et al
in Société Belge de Biochimie et de Biologie moléculaire (2002, February 22)Detailed reference viewed: 2 (2 ULg)
Cell turnover in BLV-infected sheep
; ; et al
in Aids Research and Human Retroviruses": 10th International Conference on Human Retrovirology: HTLV and Related Viruses, (2001, June)Detailed reference viewed: 7 (1 ULg)