Proteomics computational analyses suggest that hepatitis C virus E1 and pestivirus E2 envelope glycoproteins are truncated class II fusion proteins.
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Class II fusion proteins encoded by tick-borne encephalitis virus (TBEV), dengue virus, and Semliki Forest virus have a fusion peptide located at the end of a rod-like molecule comprised of three antiparallel beta sheet domains. Proteomics computational analyses suggest that hepatitis C virus (HCV) envelope glycoprotein E1 and pestivirus envelope glycoprotein E2 are truncated class II fusion proteins. Similarities were also detected between the receptor-binding portion of TBEV E and HCV E2, and between TBEV small membrane protein precursor prM and pestivirus E1. The proposed models of Flaviviridae envelope proteins can facilitate drug and vaccine development. (+info)
A histopathologic, immunohistochemical, and ultrastructural study of the intestine in pigs inoculated with classical swine fever virus.
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The aim of this study was to report on the lesions occurring in the intestine during experimental classical swine fever (CSF) and to clarify the nature of infected cells and the distribution of viral antigen. Thirty-two pigs were inoculated with the virulent CSF virus (CSFV) isolate Alfort 187 and slaughtered from 2 to 15 postinoculation days; four animals of similar background served as a control group. Immunohistochemistry, electron microscopy, and the transferase-mediated deoxyuridine triphosphate nick-end labeling method were used to detect viral antigens and apoptosis. The results showed progressive lymphoid depletion and mucosal necrosis. The lymphoid depletion could have been caused by apoptosis of lymphocytes but could not be directly attributed to the effect of CSFV on these cells. Vascular changes, pathogenic bacteria, and viral infection of epithelial cells were ruled out as causes of necrotic lesions. However, large virally infected monocytes-macrophages with ultrastructural changes indicative of activation were observed in the intestine. This suggests that monocytes-macrophages play an important role in the pathogenesis of intestinal lesions. An understanding of the function of these cells will require additional study. (+info)
Detection of tRNA-like structure through RNase P cleavage of viral internal ribosome entry site RNAs near the AUG start triplet.
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The 9600-base RNA genome of hepatitis C virus (HCV) has an internal ribosome entry site (IRES) in its first 370 bases, including the AUG start triplet at bases 342-344. Structural elements of this and other IRES domains substitute for a 5' terminal cap structure in protein synthesis. Recent work (Nadal, A., Martell, M., Lytle, J. R., Lyons, A. J., Robertson, H. D., Cabot, B., Esteban, J. I., Esteban, R., Guardia, J., and Gomez, J. (2002) J. Biol. Chem. 277, 30606-30613) has demonstrated that the host pre-tRNA processing enzyme, RNase P, can cleave the HCV RNA genome at a site in the IRES near the AUG initiator triplet. Although this step is unlikely to be part of the HCV life cycle, such a reaction could indicate the presence of a tRNA-like structure in this IRES. Because susceptibility to cleavage by mammalian RNase P is a strong indicator of tRNA-like structure, we have conducted the studies reported here to test whether such tRNA mimicry is unique to HCV or is a general property of IRES structure. We have assayed IRES domains of several viral RNA genomes: two pestiviruses related to HCV, classical swine fever virus and bovine viral diarrhea virus; and two unrelated viruses, encephalomyocarditis virus and cricket paralysis virus. We have found similarly placed RNase P cleavage sites in these IRESs. Thus a tRNA-like domain could be a general structural feature of IRESs, the first IRES structure to be identified with a functional correlate. Such tRNA-like features could be recognized by pre-existing ribosomal tRNA-binding sites as part of the IRES initiation cycle. (+info)
Classical swine fever virus interferes with cellular antiviral defense: evidence for a novel function of N(pro).
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Classical swine fever virus (CSFV) replicates efficiently in cell lines and monocytic cells, including macrophages (MPhi), without causing a cytopathic effect or inducing interferon (IFN) secretion. In the present study, the capacity of CSFV to interfere with cellular antiviral activity was investigated. When the porcine kidney cell line SK-6 was infected with CSFV, there was a 100-fold increased capacity to resist to apoptosis induced by polyinosinic-polycytidylic acid [poly(IC)], a synthetic double-stranded RNA. In MPhi, the virus infection inhibited poly(IC)-induced alpha/beta IFN (type I IFN) synthesis. This interference with cellular antiviral defense correlated with the presence of the viral N(pro) gene. Mutants lacking the N(pro) gene (DeltaN(pro) CSFV) did not protect SK-6 cells from poly(IC)-induced apoptosis, despite growth properties and protein expression levels similar to those of the wild-type virus. Furthermore, DeltaN(pro) CSFV did not prevent poly(IC)-induced type I IFN production in MPhi but rather induced type I IFN in the absence of poly(IC) in both MPhi and the porcine kidney cell line PK-15, but not in SK-6 cells. With MPhi and PK-15, an impaired replication of the DeltaN(pro) CSFV compared with wild-type virus was noted. In addition, DeltaN(pro) CSFV, but not wild-type CSFV, could interfere with vesicular stomatitis virus replication in PK-15 cells. Taken together, these results provide evidence for a novel function associated with CSFV N(pro) with respect to the inhibition of the cellular innate immune system. (+info)
Analysis of hepatitis C virus/classical swine fever virus chimeric 5'NTRs: sequences within the hepatitis C virus IRES are required for viral RNA replication.
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Hepatitis C virus (HCV) is classified in the genus Hepacivirus of the family Flaviviridae, whose members have a single-stranded RNA genome of positive polarity, which encodes a single polyprotein. Within this family, HCV is closely related to viruses of the genus Pestivirus, which includes classical swine fever virus (CSFV). Translation of the hepaci- and pestiviral polyprotein is initiated by internal entry of ribosomes, promoted by the 5'NTR. The secondary and tertiary RNA structures of the HCV and pestivirus 5'NTRs are well conserved, despite the fact that their sequences differ significantly from one another. By analogy with other positive-stranded RNA viruses, the 5'NTR of HCV is likely to contain cis-acting determinants for replication as well as the determinants for translation. Studies on both signals could be complicated, as these signals might overlap. In this study, this problem was addressed by constructing chimeric HCV/CSFV 5'NTRs. A two-step analysis of these 5'NTRs was performed: (a) in a translation assay, which provided the possibility to study translation independently of the possible effects on replication; and (b) in a replication assay, in which were studied only the chimeric 5'NTRs for which IRES-dependent translation was demonstrated. An overlap was observed between HCV RNA elements involved in these processes. Exchange of domain II had a minor effect on the translation efficiency of the chimeric 5'NTRs, while replication of subgenomic replicons with these chimeric 5'NTRs was abolished. Exchange of domain III subdomains severely decreased translation activity, while replication was maintained. (+info)
Host immune responses against hog cholera virus in pigs treated with an ionized alkali mineral complex.
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To determine the immune responses in pigs to hog cholera virus after treatment with an ionized alkali mineral complex (IAMC), 40 healthy pigs (28-32 days old) from a commercial swine farm were purchased and housed into 4 groups (n=10 each). All pigs were vaccinated intramuscularly (1 ml) with an attenuated live hog cholera virus (HCV, LOM strain) at 28-32 days old and challenged with a virulent hog cholera virus at 8 weeks after vaccination. Each group was treated with PowerFeel sprayed diet as 0.05% (w/w) in a final concentration (T-1, n=10), a diet mixed with SuperFeed as 3% (w/w) in a final concentration (T-2, n=10), or a diluted PowerFeel solution (1:500, v/v) as drinking water (T-3, n=10), respectively. A group (n=10) served as a non-treated control. Proportions of expressing CD2+ and CD8+ cells increased significantly (p<, 0.05) at 8-week post-application. Mean antibody titers of each group against HCV gradually increased to higher levels after vaccination and with challenge of the virulent virus. In conclusion, the IAMC-treated diets can be helpful for the improvement of growth in pigs with proper vaccination program, while the IAMC-treated diets have no effects on the clinical protection against hog cholera. (+info)
Humoral and cell-mediated immune responses of d/d histocompatible pigs against classical swine fever (CSF) virus.
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A better understanding of cell-mediated immune responses to classical swine fever virus (CSFV) is essential for the future development of improved vaccines. We analyzed the generation of cell-mediated and humoral immune responses in d/d histocompatible pigs following CSFV infection or vaccination. Viral infection induced high T cell responses with high primary and secondary CTL activity correlated with high IFN-gamma production, whereas vaccination with a live vaccine followed by infection mainly induced neutralizing antibody but low cell-mediated responses. Moreover, high IgG1 response was associated with high IFN-gamma response following infection whereas a weak IFN-gamma response was related to a good IgG2 response but a low IgG1 production. These data could reflect Th1/Th2-like balance of immune responses depending upon immunization protocols, which has not yet been described in the pig. T-cell responses to CSFV were evidenced by CSFV-specific CD25 upregulation on CD4-CD8+, but not on CD4+CD8- cells, which further illustrated the importance of CTL responses after infection. Our results indicated that generation of cell-mediated immune responses was much higher following intranasal/oral CSFV infection than after intramuscular vaccination, which implies that the capacity of new CSFV vaccines to induce higher T-cell responses should be considered. (+info)
Heterogeneous expression of the non-structural protein p80/p125 in cells infected with different pestiviruses.
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In order to analyse the expression of the non-structural (ns) protein p80/p125 in cells infected with different pestiviruses at the protein level, radioimmunoprecipitations with the pestivirus-specific monoclonal antibody (MAb) BVD/C16 were performed. Cell lysates infected with cytopathic (cp) and non-cytopathic (ncp) bovine viral diarrhoea (BVD) virus strains and isolates, and with hog cholera (HC) virus strains were analysed. From cpBVD virus-infected cells, the MAb precipitated one or more proteins corresponding to ns p125, displaying a marked size heterogeneity. In contrast, the lower Mr ns p80 proteins from all cpBVD virus strains and isolates analysed had identical electrophoretic motility. The ncpBVD virus strains displayed either one single band or a doublet of the p125 protein and no p80 cleavage products. The p125 proteins precipitated from HC virus-infected cells showed no size heterogeneity. The possibility is discussed that multiple recombination events, including both insertions or deletions in the genomes of ncpBVD viruses, may lead to the heterogeneous expression of the ns p125 in cpBVD virus populations. (+info)