RESUMO
Bovine gammaherpesvirus 4 (BoGHV4) is a member of the Gammaherspivirinae subfamily, Rhadinovirus genus. Its natural host is the bovine, and it is prevalent among the global cattle population. Although the complete genome of BoGHV4 has been successfully sequenced, the functions of most of its genes remain unknown. Currently, only six strains of BoGHV4, all belonging to Genotype 1, have been sequenced. This is the first report of the nearly complete genome of Argentinean BoGHV4 strains isolated from clinical cases of abortion, representing the first BoGHV4 Genotype 2 and 3 genomes described in the literature. Both Argentinean isolates presented the highest nt p-distance values, indicating a greater level of divergence. Overall, the considerable diversity observed in the complete genomes and open reading frames underscores the distinctiveness of both Argentinean isolates compared to the existing BoGHV4 genomes. These findings support previous studies that categorized the Argentinean BoGHV4 strains 07-435 and 10-154 as Genotypes 3 and 2, respectively. The inclusion of these sequences represents a significant expansion to the currently limited pool of BoGHV4 genomes while providing an important basis to increase the knowledge of local isolates.
Assuntos
Aborto Animal , Doenças dos Bovinos , Genoma Viral , Infecções por Herpesviridae , Herpesvirus Bovino 4 , Sequenciamento Completo do Genoma , Animais , Bovinos , Feminino , Gravidez , Aborto Animal/virologia , Argentina , Doenças dos Bovinos/virologia , DNA Viral/genética , Variação Genética , Genótipo , Infecções por Herpesviridae/veterinária , Infecções por Herpesviridae/virologia , Herpesvirus Bovino 4/genética , Herpesvirus Bovino 4/isolamento & purificação , Fases de Leitura Aberta , FilogeniaRESUMO
Some viruses encode inhibitory factors of apoptosis during infection to prolong cell viability and then to achieve a higher production of viral progeny or facilitate persistent infections. There is evidence that some gammaherpesviruses, including BoHV-4, carry genes that can both inhibit or induce apoptosis. BoHV-4 possesses two genes (ORF16 and ORF71) that code for proteins with anti-apoptotic functions, such as v-Bcl2 and v-Flip, respectively. Thus, it is relevant to study BoHV-4 in relation to the modulation of apoptosis in infected cells as a strategy for persistence in the host. The objective of this work was to analyze whether variations in v-Flip and v- Bcl2 of six phylogenetically divergent Argentinean isolates of BoHV-4 can influence the capacity of these strains to induce apoptosis in cell cultures. In this study, variations were mainly detected in the v-Flip gene and protein of the BoHV-4 strains belonging to genotype 3. Thus, it is possible to infer that sequence variations could be associated with some BoHV-4 genotype. Induction of apoptosis was not a significant event for any of the genetically distinct local isolates of BoHV-4 and there was not an evident relationship between the variability of both genes with the apoptotic effect of the phylogenetically distinct strains.
Assuntos
Apoptose/genética , Herpesvirus Bovino 4/genética , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Virais/genética , Sequência de Aminoácidos , Animais , Argentina , Sequência de Bases , Proteína Reguladora de Apoptosis Semelhante a CASP8 e FADD/genética , Bovinos , Doenças dos Bovinos/virologia , Linhagem Celular Tumoral , Genótipo , Células HeLa , Infecções por Herpesviridae/veterinária , Herpesvirus Bovino 4/isolamento & purificação , Humanos , Alinhamento de SequênciaRESUMO
Bovine herpevsirus 4 (BoHV-4) is a gammaherpesvirus that has been associated with different clinical conditions in cattle. In Argentina, BoHV-4 was detected in diverse bovine samples. The aim of this study was to analyze the genetic relationship of 48 field BoHV-4 strains isolated from cattle in Argentina. According to thymidine kinase (tk) gene sequences, BoHV-4 isolates belong to genotypes 1, 2 and 3. Phylogenetic analyses confirmed the presence of the three previously described viral genotypes. However, some of the studied isolates presented conflicting phylogenetic signals between the studied markers. This suggests a complex evolutionary background, that is a history of recombination, incomplete lineage sorting (deep coalescence) or a combination of these, which requires further study. These potential events make difficult the diagnosis of BoHV-4 from clinical samples of cattle and may pose a significant problem for the control of the virus in the herds.
Assuntos
Herpesvirus Bovino 4/genética , Timidina Quinase/genética , Animais , Argentina , Evolução Biológica , Bovinos/virologia , Doenças dos Bovinos/virologia , DNA Viral/genética , Evolução Molecular , Genótipo , Herpesvirus Bovino 4/isolamento & purificação , Herpesvirus Bovino 4/patogenicidade , FilogeniaRESUMO
Bovine herpesvirus 4 (BoHV-4) is increasingly believed to be responsible for several disorders of the bovine reproductive tract. The first characterization of BoHV-4 in Argentina was from samples from an aborted fetus. Argentinean isolates are highly diverse and are phylogenetically grouped in three genotypes. In this study, we describe the isolation of BoHV-4 from a bovine fetus with a gestational age of 8 months and without macroscopic lesions. Genetic analyses revealed that the isolated strain belongs to genotype 2. This is the first report on the presence of infectious BoHV-4 in tissues from an aborted bovine fetus.
Assuntos
Feto Abortado/virologia , Aborto Animal/virologia , Doenças dos Bovinos/virologia , Infecções por Herpesviridae/veterinária , Herpesvirus Bovino 4/genética , Herpesvirus Bovino 4/isolamento & purificação , Aborto Animal/epidemiologia , Animais , Argentina/epidemiologia , Sequência de Bases , Bovinos , Doenças dos Bovinos/epidemiologia , Infecções por Herpesviridae/epidemiologia , Infecções por Herpesviridae/virologia , RNA Viral/genéticaRESUMO
Bovine gammaherpesvirus 4 (BoHV4) is a member of the family Herpesviridae. In Argentina, BoHV4 was isolated and characterized in 2007 from samples of aborted cows. Argentinean isolates are highly divergent and are classified as: Genotype 1(Movar-like), Genotype 2 (DN599-like) and Genotype 3 (a novel group). The aim of this study was to comparatively evaluate the biological characteristics of six Argentinean BoHV4 field isolates in cell lines from different origins. All strains induced productive infection in the cell lines used, with different degrees of permissiveness. A direct relationship among the times of appearance of cytopathic effect, the growth kinetics, the size of the lysis plaques and the virulent-like behaviour in vitro could not be established. However, although slight, there are differences in the biological behaviour of the BoHV4 fields isolates analyzed. This variability is independent of their genetic classification but would be conditioned by the nature of the infected cells.
Assuntos
Infecções por Herpesviridae/veterinária , Herpesvirus Bovino 4/fisiologia , Infecções Tumorais por Vírus/veterinária , Replicação Viral/fisiologia , Animais , Argentina , Bovinos , Linhagem Celular , Chlorocebus aethiops , Efeito Citopatogênico Viral , Cães , Células HeLa , Células Hep G2 , Herpesvirus Bovino 4/genética , Herpesvirus Bovino 4/isolamento & purificação , Humanos , Células Madin Darby de Rim Canino , Células VeroRESUMO
Bovine herpesvirus 4 (BoHV-4) is a gammaherpesvirus, belonging to the Rhadinovirus genus, which is increasingly associated with various problems of the reproductive tract of cattle. In Argentina, analysis of BoHV-4 strains isolated from cervico-vaginal mucus of aborted cows revealed a high genetic divergence among strains, which could be classified in three different groups: Genotype 1 comprises Movar-like strains (European prototype), Genotype 2 includes DN599-like strains (American prototype) and Genotype 3 corresponds to a novel genotype group. Understanding the replication behavior in cell cultures and the molecular characteristics of this pathogen of cattle is critical for the rational design of in vitro experiments. The aim of this work was to quantitatively evaluate the replication properties of different Argentinean BoHV-4 strains and to characterize their phylogenetic relationships. Significant differences were evident among the virus titers of the different BoHV-4 isolates in vitro. The most conserved gene was the major capsid protein (ORF25). The glycoprotein B (gB), glycoprotein H (gH), and thymidine kinsase (TK) genes displayed both synonymous and non-synonymous substitutions, with the highest diversity observed for gB, which displayed amino acid substitutions in 24 out of the 178 positions examined. Strains 09/759, 12/512, and 07/568 presented a deletion encompassing amino acid position 27 to 35, whereas strains 07/435 and 09/227 had a deletion from position 28 to 35. Two strains, 07/435 and 09/227, also displayed the highest divergence compared to the other strains analyzed. This study provides information about the in vitro replication and behavior of nine field isolates of BoHV-4. These findings are relevant since available information on the in vitro growth characteristics of BoHV-4 strains is scarce. The results from this study may also be useful for establishing comparisons with other related viruses.
Assuntos
Herpesvirus Bovino 4/isolamento & purificação , Herpesvirus Bovino 4/fisiologia , Replicação Viral/genética , Animais , Argentina , Bovinos , Doenças dos Bovinos/virologia , Linhagem Celular , DNA Viral/genética , Feminino , Variação Genética , Genótipo , Infecções por Herpesviridae/veterinária , Infecções por Herpesviridae/virologia , Herpesvirus Bovino 4/genética , Filogenia , Timidina Quinase/genética , Vagina/virologia , Esfregaço Vaginal/veterinária , Proteínas do Envelope Viral/genéticaRESUMO
Bovine herpesvirus 4 (BoHV-4) is increasingly considered as responsible for various problems of the reproductive tract. The virus infects mainly blood mononuclear cells and displays specific tropism for vascular endothelia, reproductive and fetal tissues. Epidemiological studies suggest its impact on reproductive performance, and its presence in various sites in the reproductive tract highlights its potential transmission in transfer-stage embryos. This work describes the biological and genetic characterization of BoHV-4 strains isolated from an in vitro bovine embryo production system. BoHV-4 strains were isolated in 2011 and 2013 from granulosa cells and bovine oocytes from ovary batches collected at a local abattoir, used as "starting material" for in vitro production of bovine embryos. Compatible BoHV-4-CPE was observed in the co-culture of granulosa cells and oocytes with MDBK cells. The identity of the isolates was confirmed by PCR assays targeting three ORFs of the viral genome. The phylogenetic analyses of the strains suggest that they were evolutionary unlinked. Therefore it is possible that BoHV-4 ovary infections occurred regularly along the evolution of the virus, at least in Argentina, which can have implications in the systems of in vitro embryo production. Thus, although BoHV-4 does not appear to be a frequent risk factor for in vitro embryo production, data are still limited. This study reveals the potential of BoHV-4 transmission via embryo transfer. Moreover, the high variability among the BoHV-4 strains isolated from aborted cows in Argentina highlights the importance of further research on the role of this virus as an agent with the potential to cause reproductive disease in cattle. The genetic characterization of the isolated strains provides data to better understand the pathogenesis of BoHV-4 infections. Furthermore, it will lead to fundamental insights into the molecular aspects of the virus and the means by which these strains circulate in the herds.
Assuntos
Embrião de Mamíferos/virologia , Células da Granulosa/virologia , Herpesvirus Bovino 4/genética , Oócitos/virologia , Animais , Argentina , Teorema de Bayes , Bovinos , Células Cultivadas , Técnicas de Cocultura , DNA Viral/análise , Cães , Feminino , Células da Granulosa/citologia , Herpesvirus Bovino 4/classificação , Herpesvirus Bovino 4/isolamento & purificação , Células Madin Darby de Rim Canino , Oócitos/citologia , Fases de Leitura Aberta/genética , Filogenia , Análise de Sequência de DNARESUMO
Bovine Herpesvirus 4 (BoHV-4) is a member of Gammaherpesvirinae sub-family and belongs to genus Rhadinovirus . This virus has been associated with different clinical manifestations and research activity has put forward a strong correlation among virus infection, postpartum metritis, and abortion. The goal of this work was to characterize a virus strain isolate from a cow's uterine outflow. From swabs drawn of uterine secretion, a virus strain was isolated and characterized by its cytopathology, morphology, and molecular biology approaches. In culture there was CPE development, characterized mainly by long strands with several small balloons along them, radiated from infected cells. Electron microscopy analysis revealed virus particles that had icosahedrical capsid symmetry surrounded by a loose envelope, typical of a herpesvirus. A 2,571 bp PCR product after Hind III digestion generated four fragments, whose base pair composition were 403, 420, 535, and 1,125 bp. Restriction enzymes Hind III and Bam HI generated the expected diagnostic bands as well as a 2,350 bp hypermolar fragment as a result of Bam HI treatment to demonstrate that agent was a bovine herpesvirus 4, appertaining to DN-599 group.
Assuntos
Doenças dos Bovinos/virologia , Infecções por Herpesviridae/veterinária , Herpesvirus Bovino 4/classificação , Herpesvirus Bovino 4/isolamento & purificação , Infecções Tumorais por Vírus/veterinária , Animais , Brasil , Bovinos , Efeito Citopatogênico Viral , DNA Viral/genética , DNA Viral/metabolismo , Exsudatos e Transudatos/virologia , Feminino , Infecções por Herpesviridae/virologia , Herpesvirus Bovino 4/genética , Microscopia Eletrônica de Transmissão , Polimorfismo de Fragmento de Restrição , Infecções Tumorais por Vírus/virologia , Útero/patologia , Útero/virologia , Vírion/ultraestrutura , Cultura de VírusRESUMO
Bovine herpesvirus 4 (BoHV-4) has been isolated from cattle with respiratory infections, vulvovaginitis, mastitis, abortions, endometritis and from apparently healthy animals throughout the world. Although it has not yet been established as causal agent of a specific disease entity, it is primarily associated with reproductive disorders of cattle. This virus can infect a wide range of species, either in vivo or in vitro. Two groups of prototype strains were originated from the first isolates: the DN599-type strains (American group) and the Movar-type strains (European group). In Argentina, BoHV-4 was isolated and characterized in 2007 from vaginal discharge samples taken from cows that had aborted. So far, more than 40 isolates, mainly associated with aborting bovine females have been registered in our country.
Assuntos
Aborto Animal/virologia , Doenças dos Bovinos/virologia , Infecções por Herpesviridae/veterinária , Herpesvirus Bovino 4/isolamento & purificação , Infecções Tumorais por Vírus/veterinária , Aborto Animal/epidemiologia , Animais , Anticorpos Antivirais/sangue , Apoptose , Argentina/epidemiologia , Bovinos , Doenças dos Bovinos/epidemiologia , Causalidade , Efeito Citopatogênico Viral , Endométrio/virologia , Feminino , Genoma Viral , Infecções por Herpesviridae/diagnóstico , Infecções por Herpesviridae/epidemiologia , Infecções por Herpesviridae/virologia , Herpesvirus Bovino 4/classificação , Herpesvirus Bovino 4/patogenicidade , Herpesvirus Bovino 4/fisiologia , Especificidade de Hospedeiro , Interações Hospedeiro-Patógeno , Transtornos Puerperais/veterinária , Transtornos Puerperais/virologia , Infecções Tumorais por Vírus/diagnóstico , Infecções Tumorais por Vírus/epidemiologia , Infecções Tumorais por Vírus/virologia , Tropismo Viral , Virulência , Ativação ViralRESUMO
Bovine Herpesvirus 4 (BoHV-4) is a member of Gammaherpesvirinae sub-family and belongs to genus Rhadinovirus. This virus has been associated with different clinical manifestations and research activity has put forward a strong correlation among virus infection, postpartum metritis, and abortion. The goal of this work was to characterize a virus strain isolate from a cows uterine outflow. From swabs drawn of uterine secretion, a virus strain was isolated and characterized by its cytopathology, morphology, and molecular biology approaches. In culture there was CPE development, characterized mainly by long strands with several small balloons along them, radiated from infected cells. Electron microscopy analysis revealed virus particles that had icosahedrical capsid symmetry surrounded by a loose envelope, typical of a herpesvirus. A 2,571 bp PCR product after HindIII digestion generated four fragments, whose base pair composition were 403, 420, 535, and 1,125 bp. Restriction enzymes HindIII and BamHI generated the expected diagnostic bands as well as a 2,350 bp hypermolar fragment as a result of BamHI treatment to demonstrate that agent was a bovine herpesvirus 4, appertaining to DN-599 group.(AU)
Assuntos
Animais , Feminino , Bovinos , Doenças dos Bovinos/virologia , Infecções por Herpesviridae/veterinária , Herpesvirus Bovino 4/classificação , Herpesvirus Bovino 4/isolamento & purificação , Infecções Tumorais por Vírus/veterinária , Brasil , Efeito Citopatogênico Viral , DNA Viral/genética , DNA Viral/metabolismo , Exsudatos e Transudatos/virologia , Infecções por Herpesviridae/virologia , Herpesvirus Bovino 4/genética , Microscopia Eletrônica de Transmissão , Polimorfismo de Fragmento de Restrição , Útero/patologia , Útero/virologia , Vírion/ultraestrutura , Cultura de VírusRESUMO
Bovine herpesvirus 4 (BoHV-4) is a γ-herpesvirus that has been isolated both from apparently healthy animals and from cattle with a variety of clinical signs, including post-partum endometritis and abortion. BoHV-4 causes either a persistent or a latent infection in cells of the monocyte/macrophage lineage. Two groups of BoVH-4 strains have been defined based on their restriction patterns: the Movar-like strains (European prototype) and the DN 599-like strains (American prototype). The purpose of the present study was to genetically characterize wild type BoHV-4 strains isolated from vaginal discharges of aborted cows in Argentina. The virus was identified by isolation and nested PCR in all vaginal discharge samples from aborted cows, either as a sole agent or in association with other pathogens. Restriction enzyme profiling and phylogenetic analysis demonstrated that there is a high genetic variability among the studied field isolates. The existence of three groups of strains, which were designated as genotypes 1, 2 and 3, is described. Genotypes 1 and 2 possibly correspond to the Movar-like and DN 599-like groups, respectively, whereas Genotype 3 corresponds to a novel group. Two viral strains did not cluster into any of these three groups, indicating that other genotypes could be circulating in Argentina. These results suggest a complex epidemiological background for the Argentinean BoHV-4 strains, probably influenced by independent events of genetic drift. This hypothesis cannot be rejected based on the available data. However, there is no direct evidence supporting this possibility. Thus, it seems speculative to suggest that interspecific jumps are responsible for the observed phylogenetic grouping. On the other hand, our analyses suggest a geographical structure for the observed viral genotypes, since genotypes 1 and 2 included the European (Movar-like) and American (DN599-like) reference strains, respectively. Geographic dispersion is known to be a driver of herpes viruses diversification, and independent evolution in geographical isolated places ensures the emergence of particular mutations in each location due to genetic drift (Compans, 2007; Zong et al., 1999). Therefore, at this point, the genetic drift hypothesis is the one that requires less ad-hoc considerations and thus, to our understanding, is the one that fits to the findings from this study. The involvement of this genetic variability in the detection and pathogenesis of BoHV-4 remains to be investigated.