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1.
BMC Genomics ; 20(1): 516, 2019 Jun 21.
Artigo em Inglês | MEDLINE | ID: mdl-31226933

RESUMO

BACKGROUND: Bovine viral diarrhea virus (BVDV) is an economically important viral pathogen of domestic and wild ruminants. Apart from cattle, small ruminants (goats and sheep) are also the susceptible hosts for BVDV. BVDV infection could interfere both of the innate and adaptive immunity of the host, while the genes and mechanisms responsible for these effects have not yet been fully understood. Peripheral blood mononuclear cells (PBMCs) play a pivotal role in the immune responses to viral infection, and these cells were the target of BVDV infection. In the present study, the transcriptome of goat peripheral blood mononuclear cells (PBMCs) infected with BVDV-2 was explored by using RNA-Seq technology. RESULTS: Goat PBMCs were successfully infected by BVDV-2, as determined by RT-PCR and quantitative real-time RT-PCR (qRT-PCR). RNA-Seq analysis results at 12 h post-infection (hpi) revealed 499 differentially expressed genes (DEGs, fold-change ≥ ± 2, p < 0.05) between infected and mock-infected PBMCs. Of these genes, 97 were up-regulated and the remaining 352 genes were down-regulated. The identified DEGs were found to be significantly enriched for locomotion/ localization, immune response, inflammatory response, defense response, regulation of cytokine production, etc., under GO enrichment analysis. Cytokine-cytokine receptor interaction, TNF signaling pathway, chemokine signaling pathway, etc., were found to be significantly enriched in KEGG pathway database. Protein-protein interaction (PPI) network analysis indicated most of the DEGs related to innate or adaptive immune responses, inflammatory response, and cytokine/chemokine-mediated signaling pathway. TNF, IL-6, IL-10, IL-12B, GM-CSF, ICAM1, EDN1, CCL5, CCL20, CXCL10, CCL2, MAPK11, MAPK13, CSF1R and LRRK1 were located in the core of the network and highly connected with other DGEs. CONCLUSIONS: BVDV-2 infection of goat PBMCs causes the transcription changes of a series of DEGs related to host immune responses, including inflammation, defense response, cell locomotion, cytokine/chemokine-mediated signaling, etc. The results will be useful for exploring and further understanding the host responses to BVDV-2 infection in goats.


Assuntos
Vírus da Diarreia Viral Bovina Tipo 2/genética , Doenças das Cabras/imunologia , Doenças das Cabras/virologia , Infecções por Pestivirus/veterinária , Animais , Vírus da Diarreia Viral Bovina Tipo 2/imunologia , Perfilação da Expressão Gênica , Regulação Viral da Expressão Gênica , Doenças das Cabras/genética , Cabras , Imunidade/genética , Leucócitos Mononucleares/virologia , Infecções por Pestivirus/genética , Infecções por Pestivirus/imunologia , Análise de Sequência de RNA , Replicação Viral
2.
BMC Vet Res ; 15(1): 202, 2019 Jun 14.
Artigo em Inglês | MEDLINE | ID: mdl-31200717

RESUMO

BACKGROUND: Bovine viral diarrhea virus (BVDV) infects various ungulates and causes reproductive failure in infected goats. BVDV has been detected among goats in the Republic of Korea, but the route of transmission remains unclear. Here, we aimed to investigate whether BVDV-1b circulating among Korean cattle can be transmitted to Korean native goats (Capra aegagrus hircus) and characterize the outcomes of BVDV infection in these goats. RESULTS: Four goats were inoculated intranasally with the Korean noncytopathic (ncp) BVDV-1b strain. Two goats exhibited clinical signs of illness, including coughing and nasal discharge. Nasal swabs and blood were collected to screen for viral RNA and BVDV antibodies. Using the 5'-untranslated region (UTR), viral RNA was detected in the nasal swabs of two goats (Goat 1 and 3) on 12 day post-inoculation (dpi) and in the blood sample of one goat (Goat 1) on 7 and 19 dpi. Using the N-terminal protease (Npro) region, viral RNA was detected in the blood sample of Goat 1 on 7 and 12 dpi. Antibodies to BVDV were detected in Goats 1 and 3 on 16-21 dpi using enzyme-linked immunosorbent assay. Sequence analysis of the virus from nasal swabs and blood samples, which was detected via RT-PCR, using the 5'-UTR and Npro regions led to the identification of the strain as ncp BVDV-1b and revealed changes in the nucleotide sequence of these goats. CONCLUSIONS: Our results indicate that changes in the nucleotide sequence are associated with the establishment of BVDV infection in Korean native goats; these changes may be owing to a process required for the establishment of infection in a new host reservoir. Broadly, these findings highlight the importance of BVDV surveillance in ungulates other than cattle.


Assuntos
Vírus da Diarreia Viral Bovina Tipo 1/genética , Doenças das Cabras/virologia , Infecções por Pestivirus/veterinária , Regiões 5' não Traduzidas , Animais , Anticorpos Antivirais/sangue , Vírus da Diarreia Viral Bovina Tipo 1/imunologia , Vírus da Diarreia Viral Bovina Tipo 1/patogenicidade , Ensaio de Imunoadsorção Enzimática/veterinária , Feminino , Doenças das Cabras/transmissão , Cabras , Mucosa Nasal/virologia , Infecções por Pestivirus/genética , Infecções por Pestivirus/virologia , RNA Viral/sangue , RNA Viral/genética , República da Coreia , Análise de Sequência de RNA
3.
Virus Genes ; 55(3): 298-303, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30706196

RESUMO

Bungowannah virus, which belongs to the genus Pestivirus within the family Flaviviridae, has been associated with myocarditis and a high incidence of stillbirths in pigs. In 2003, the virus was initially detected in a large pig farming complex on two separate sites in New South Wales, Australia. Until now, it has not been detected at other locations. Despite a program of depopulation and disinfection, the virus could be only eradicated from one of the affected farm complexes, the Bungowannah unit, but became endemic on the second complex, the Corowa unit. In the present study, the genetic variability of virus isolates collected between 2003 and 2014 in the endemically infected population has been retrospectively investigated. Phylogenetic analysis carried out based on sequences of the E2 and NS5B coding regions and the full-length open-reading frame revealed that the isolates from the different farm sites are closely related, but that samples collected between 2010 and 2014 at the Corowa farm site clustered in a different branch of the phylogenetic tree. Since 2010, a high-genetic stability of this RNA virus within the Corowa farm complex, probably due to an effective adaptation of the virus to the affected pig population, could be observed.


Assuntos
Infecções por Pestivirus/genética , Pestivirus/genética , Natimorto/genética , Doenças dos Suínos/genética , Animais , Austrália , Surtos de Doenças , Pestivirus/patogenicidade , Infecções por Pestivirus/veterinária , Infecções por Pestivirus/virologia , Estudos Retrospectivos , Natimorto/veterinária , Suínos , Doenças dos Suínos/virologia
4.
Sci Rep ; 7: 44459, 2017 03 14.
Artigo em Inglês | MEDLINE | ID: mdl-28290554

RESUMO

A multitude of viral factors - either inhibiting the induction of the IFN-system or its effectors - have been described to date. However, little is known about the role of structural components of the incoming virus particle in protecting against IFN-induced antiviral factors during or immediately after entry. In this study, we take advantage of the previously reported property of Classical swine fever virus (family Flaviviridae, genus Pestivirus) to tolerate a deletion of the core protein if a compensatory mutation is present in the NS3-helicase-domain (Vp447∆c). In contrast to the parental virus (Vp447), which causes a hemorrhagic-fever-like disease in pigs, Vp447∆c is avirulent in vivo. In comparison to Vp447, growth of Vp447∆c in primary porcine cells and IFN-treated porcine cell lines was reduced >20-fold. Also, primary porcine endothelial cells and IFN-pretreated porcine cell lines were 8-24 times less susceptible to Vp447∆c. This reduction of susceptibility could be partially reversed by loading Vp447∆c particles with different levels of core protein. In contrast, expression of core protein in the recipient cell did not have any beneficial effect. Therefore, a protective effect of core protein in the incoming virus particle against the products of IFN-stimulated genes could be demonstrated.


Assuntos
Vírus da Febre Suína Clássica/genética , Infecções por Pestivirus/genética , Pestivirus/genética , Proteínas do Core Viral/genética , Animais , Vírus da Febre Suína Clássica/patogenicidade , Vírus de DNA/genética , Pestivirus/patogenicidade , Infecções por Pestivirus/virologia , Suínos/virologia , Replicação Viral/genética
5.
Virus Genes ; 53(2): 233-239, 2017 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-27866318

RESUMO

In this study, we constructed for the first time a full-length cDNA clone of pig-original bovine viral diarrhea virus 2 (BVDV-2) strain SH-28, modified the cDNA clone (pASH28) for mutant pASHΔNpro and derived virus strain vASHΔNpro by deleting the genomic region encoding the Npro polypeptide, and examined significance of protein Npro for antiviral responses in vitro. Data showed that Npro-deletion mutant virus vASHΔNpro led to significant overexpression of oligo adenylate synthetase (OAS), myxovirus-resistant protein 1 (Mx1), and ubiquitin-like protein 15 (ISG15). Data also revealed that overexpression of Npro, but not NS2 and NS3 proteins, resulted in significant down-regulation of OAS, Mx1, and ISG15 production (p ≤ 0.05) in bovine cells as well as porcine cells transfected with Npro recombinant eukaryotic expression plasmids. Npro (but not NS2 and NS3) was also found to inhibit poly(IC) from inducing production of type I interferon (IFN-I). These results indicated that protein Npro may play multiple roles in regulating antiviral response in host cells interfered by pig BVDV-2 strain, and provided useful information to understand better the mechanism of BVDV-2 persistent infection in pigs.


Assuntos
Vírus da Diarreia Viral Bovina Tipo 2/genética , Proteínas de Resistência a Myxovirus/genética , Infecções por Pestivirus/genética , Proteínas não Estruturais Virais/genética , Animais , Bovinos , Linhagem Celular , Vírus da Diarreia Viral Bovina Tipo 2/patogenicidade , Regulação Viral da Expressão Gênica , Proteínas de Resistência a Myxovirus/biossíntese , Infecções por Pestivirus/virologia , Suínos/virologia , Ubiquitinas/biossíntese , Ubiquitinas/genética , Proteínas não Estruturais Virais/biossíntese
6.
Vet Microbiol ; 168(2-4): 312-7, 2014 Jan 31.
Artigo em Inglês | MEDLINE | ID: mdl-24388633

RESUMO

Llamas and alpacas are domesticated South American camelids (SACs) important to ancestral population in the Altiplano region, and to different communities worldwide where they have been introduced. These ungulates have shown to be susceptible to several livestock viral pathogens such as members of the Pestivirus genus, in particular Bovine Viral Diarrhea (BVDV), but there is little data available on Pestivirus infections in SACs. In this study we aimed to detect and identify Pestivirus genotypes and subgroups infecting SACs in both wild and confined environments. Samples were collected from 136 llamas and 30 alpacas from different areas in the Chilean Altiplano (wild animals), and from 22 llamas and 26 alpacas diagnosed as Pestivirus positive from the Metropolitana region in Chile (confined animals). Seroneutralization tests showed titers lower than 2 in all 166 samples from Chilean Altiplano. These samples were also negative to BVDV isolation, indicating that these animals have not been exposed to Pestivirus. After reactivation of positive samples from the Metropolitana region, the 5' non-codifying region (5'NCR) and E2 glycoprotein were amplified by RT-PCR from the Pestivirus genome. Viral sequences were pairwise compared and phylogenetic trees were constructed. The 5'NCR analysis showed that all 12 sequenced isolates belonged to BVDV-1. Of particular interest, isolates from eight llama and two alpaca were BVDV-1j and two alpacas were BVDV-1b. In agreement with these results, E2 phylogenetic analysis rendered a similar grouping indicating that all 16 isolates belong to BVDV-1. However, the lower availability of E2 sequences determines the creation of a smaller number of sub-groups than the 5'NCR sequences. Based on the E2 sequences, the 5'NCR BVDV 1j group consisting of all the llamas and 3 alpacas are completely included in the E2 BVDV 1e group. Due to the universal availability of the 5'NCR segment, we propose the classification of these Chilean llamas and alpacas Pestivirus isolates as BVDV 1j and BVDV 1b respectively. Thus, this is the first time BVDV-1j is obtained in SACs. In addition, these results indicate Pestivirus infection in llamas and alpacas is associated with bovine population as genotypes and sub-groups are the same as those affecting Chilean livestock.


Assuntos
Camelídeos Americanos/virologia , Vírus da Diarreia Viral Bovina Tipo 1/genética , Infecções por Pestivirus/veterinária , Infecções por Pestivirus/virologia , Animais , Chile , Vírus da Diarreia Viral Bovina Tipo 1/classificação , Vírus da Diarreia Viral Bovina Tipo 1/isolamento & purificação , Genótipo , Infecções por Pestivirus/genética , Filogenia , Reação em Cadeia da Polimerase Via Transcriptase Reversa
7.
Vet Microbiol ; 139(3-4): 245-52, 2009 Nov 18.
Artigo em Inglês | MEDLINE | ID: mdl-19577384

RESUMO

Classical swine fever is a highly contagious viral disease causing severe economic losses in pig production almost worldwide. All pestivirus species can infect pigs, therefore accurate and rapid pestivirus detection and differentiation is of great importance to assure control measures in swine farming. Here we describe the development and evaluation of a novel multiplex, highly sensitive and specific RT-PCR for the simultaneous detection and rapid differentiation between CSFV and other pestivirus infections in swine. The universal and differential detection was based on primers designed to amplify a fragment of the 5' non-coding genome region for the detection of pestiviruses and a fragment of the NS5B gene for the detection of classical swine fever virus. The assay proved to be specific when different pestivirus strains from swine and ruminants were evaluated. The analytical sensitivity was estimated to be as little as 0.89TCID(50). The assay analysis of 30 tissue homogenate samples from naturally infected and non-CSF infected animals and 40 standard serum samples evaluated as part of two European Inter-laboratory Comparison Tests conducted by the European Community Reference Laboratory, Hanover, Germany proved that the multiplex RT-PCR method provides a rapid, highly sensitive, and cost-effective laboratory diagnosis for classical swine fever and other pestivirus infections in swine.


Assuntos
Peste Suína Clássica/diagnóstico , Infecções por Pestivirus/diagnóstico , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Animais , Peste Suína Clássica/genética , Peste Suína Clássica/virologia , Vírus da Febre Suína Clássica/genética , Primers do DNA/análise , Primers do DNA/genética , Diagnóstico Diferencial , Pestivirus/genética , Infecções por Pestivirus/genética , Sensibilidade e Especificidade , Sus scrofa
8.
J Virol ; 81(7): 3327-38, 2007 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-17215285

RESUMO

Different genetically engineered mutants of bovine viral diarrhea virus (BVDV) were analyzed for the ability to establish infection in the fetuses of pregnant heifers. The virus mutants exhibited either a deletion of the overwhelming part of the genomic region coding for the N-terminal protease N(pro), a deletion of codon 349, which abrogates the RNase activity of the structural glycoprotein E(rns), or a combination of both mutations. Two months after infection of pregnant cattle with wild-type virus or either of the single mutants, the majority of the fetuses contained virus or were aborted or found dead in the uterus. In contrast, the double mutant was not recovered from fetal tissues after a similar challenge, and no dead fetuses were found. This result was verified with a nonrelated BVDV containing similar mutations. After intrauterine challenge with wild-type virus, mutated viruses, and cytopathogenic BVDV, all viruses could be detected in fetal tissue after 5, 7, and 14 days. Type 1 interferon (IFN) could be detected in fetal serum after challenge, except with wild-type noncytopathogenic BVDV. On days 7 and 14 after challenge, the largest quantities of IFN in fetal serum were induced by the N(pro) and RNase-negative double mutant virus. The longer duration of fetal infection with the double mutant resulted in abortion. Therefore, for the first time, we have demonstrated the essential role of both N(pro) and E(rns) RNase in blocking interferon induction and establishing persistent infection by a pestivirus in the natural host.


Assuntos
Vírus da Diarreia Viral Bovina Tipo 1/fisiologia , Feto/enzimologia , Feto/virologia , Ribonucleases/metabolismo , Proteínas Virais/metabolismo , Animais , Bovinos , Linhagem Celular , DNA Complementar/genética , Cães , Feto/embriologia , Deleção de Genes , Genoma/genética , Interferons/metabolismo , Dados de Sequência Molecular , Mutação/genética , Infecções por Pestivirus/enzimologia , Infecções por Pestivirus/genética , Infecções por Pestivirus/prevenção & controle , Infecções por Pestivirus/virologia , Ribonucleases/genética , Fatores de Tempo , Proteínas Virais/genética
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