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ABSTRACT: Avian tuberculosis was diagnosed via histopathology, microbiology, and molecular biology in two of six pheasants from a local sanctuary bird house in Taiwan. Swinehoe's pheasant (Lophura swinhoii) is a near-threatened species in Taiwan. The infected birds showed clinical signs such as fatigue, inappetence, diarrhea, and fluffing of feathers. On postmortem, nonmineralized caseogranulomas were present in the brain, heart, lung, liver, spleen, costal membranes, and intestinal tracts. The presence of granulomas in the lungs of the infected pheasants may suggest that exposure to the infective agent was via the respiratory route rather than the alimentary route. Histopathologic findings were typical of avian tuberculosis, including acid-fast bacilli and centrally located caseous necrosis surrounded by epitheloid macrophages, lymphocytes, and multinucleated giant cells. Laboratory confirmation was made based on lesions and via Ziehl-Neelsen acid-fast stain, polymerase chain reaction, nucleic acid sequencing, and a reliable assay protocol for identification of diseases bioactive amplification with probing assay.
Avian Diseases 12/2009; 53(4):629-33. · 1.46 Impact Factor
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ABSTRACT: Although induction of apoptosis by bovine ephemeral fever virus (BEFV) in several cell lines has been previously demonstrated by our laboratory, less information is available on the process of BEFV-induced apoptosis in terms of cellular pathways and specific proteins involved. In order to determine the step in viral life cycle at which apoptosis of infected cells is triggered, chemical and physical agents were used to block viral infection. Treatment of BHK-21 infected cells with ammonium chloride (NH4Cl) or cells infected with UV-inactivated BEFV was seen to abrogate virus apoptosis induction, suggesting that virus uncoating and gene expression are required for the induction of apoptosis. Using soluble death receptors Fc:Fas chimera to block Fas signaling, BEFV-induced apoptosis was inhibited in cells. BEFV infection of BHK-21 cells results in the Fas-dependent activation of caspase 8 and cleavage of Bid. This initiated the dissipation of the membrane potential and the release of cytochrome c but not AIF or Smac/DIABLO from mitochondrial into cytoplasm leading to activation of caspase 9. Combined activation of the death receptor and mitochondrial pathways results in activation of the downstream effecter caspase 3 leading to cleavage of PARP. Fas-mediated BEFV-induced apoptosis could be suppressed by the overexpression of Bcl-2 or by treatment with caspase inhibitors and soluble death receptors Fc:Fas chimera. Taken together, this study provided first evidence demonstrating that BEFV-induced apoptosis requires viral gene expression and occurs through the activation of Fas and mitochondrion-mediated caspase-dependent pathways.
Apoptosis 08/2009; 14(7):864-77. · 4.07 Impact Factor
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Apoptosis 07/2009; · 4.07 Impact Factor
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ABSTRACT: Classical swine fever virus (CSFV) causes significant losses in the pig industry in many countries. NS3 proteins of CSFV, which include serine protease and RNA helicase/nucleotide triphosphatase (NTPase) activities, are multifunctional proteins involved in polyprotein processing and viral replication. Previous reports showed that NS3 protein can induce apoptosis in host cells that present cytopathic effects (CPE). Baculovirus/insect cell systems are used widely for recombinant protein production. In this study, one recombinant baculovirus BacSC-NS3 expressing histidine-tagged NS3 with the transmembrane domain (TM) and cytoplasmic domain (CTD) derived from baculovirus envelope protein gp64 of baculovirus was constructed. After infection, NS3 was expressed and anchored to the plasma membrane of Sf-9 cells, as demonstrated by Western blot assay and confocal microscopy. Immunogold electron microscopy demonstrated that the NS3 glycoprotein successfully displayed on the baculoviral envelope. Animal vaccine tests showed that recombinant baculovirus BacSC-NS3 elicited significantly higher NS3 antibody titers in the treated mouse models than the control group. This report demonstrated the potential of NS3-pseudotyped baculovirus expression of NS3 protein successfully.
Journal of virological methods 06/2009; 159(2):259-64. · 2.13 Impact Factor
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ABSTRACT: The surface glycoprotein G is considered as the major neutralizing and protective antigen of bovine ephemeral fever virus (BEFV). Comparison of the deduced amino acid sequence of G protein of BEFV isolates during the period 1984-2004 outbreaks in Taiwan showed amino acid substitutions in the neutralizing epitopes. All the isolates differ markedly in the neutralizing epitope at the same amino acid positions compared to the currently available killed vaccine strain (Tn73). Tn88128 strain isolated in 1999 showed the maximum variability of 12 amino acids, 5 amino acid in the neutralization epitope and 7 apart from, respectively. Combinations of both Tn88128 (1999) and commercially available vaccine strain (Tn73) were developed and its safety was evaluated in mice, guinea pigs, calves, and pregnant cows. None of the animals showed any adverse effect or clinical signs. Calves were immunized with commercial vaccine (Tn73) and, combined vaccine (Tn73 and Tn88128), respectively, with adjuvants such as Al-gel and water-in-oil-in-water (w/o/w) oil and PBS alone and challenged with Tn88128 strains. Except PBS administered animals, all the vaccinated animals showed protective immune response. However, animals immunized with combined vaccine plus w/o/w adjuvant elicited stronger neutralization antibodies and long lasting immunity compared to other vaccines.
Journal of Veterinary Medical Science 07/2006; 68(6):543-8. · 0.85 Impact Factor
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ABSTRACT: A rapid, sensitive, and specific assay, RAPID-BAP assay, was developed to detect and quantify the G protein-encoding gene of bovine ephemeral fever virus (BEFV). This new technique uses a nested PCR and magnetic bead-based DNA probing assay. The optimal conditions for the assay were examined. By applying a nested PCR, a minimum of 1 copy/mul of the BEFV plasmid DNA could be detected by the assay. The optimal hybridization conditions at 50 degrees C in 5x SSC and 0.5% SDS with a 20-min incubation allowed clear discrimination between negative and positive controls. The assay was also highly specific as all negative controls failed to show any positive detection. The diagnostic sensitivity of the RAPID-BAP assay, real-time RT-PCR, and conventional RT-PCR in the detection of 34 clinical blood samples suspected to have BEFV infections were 72.73, 36.36, and 18.18%, respectively. The results indicated that the RAPID-BAP assay developed in this study was more sensitive than the conventional RT-PCR and real-time RT-PCR assays for the detection of BEFV. The novel RAPID-BAP assay is an excellent diagnostic tool with high sensitivity, specificity, and fast turnaround time.
Journal of Virological Methods 11/2005; 129(1):75-82. · 2.01 Impact Factor
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ABSTRACT: Bovine ephemeral fever (BEF), a vector-borne disease of cattle, is caused by the Ephemerovirus of the family Rhabdoviridae. In the past 40 years, Taiwan has had seven BEF epizootics, and we have previously reported the first five. This study summarizes the 2001 and 2002 epizootics; conducted case-control serologic studies on 10 herds involved in the 2001 epizootic; determined whether the recent BEF viruses have varied significantly; and discusses the relationship between epizootic patterns and possible variant BEF viruses. For mature cows that had received at least 2 doses of vaccine before the study, a negative correlation between the prevaccinated (the 3rd dose and after) serum neutralization antibody (SNA) titers and their postvaccinated peak rates was found. When prevaccinated SNA levels were at < or = 32, their postvaccinated SNA levels increased significantly faster (P<0.01) than for those at > or = 32. The glycoprotein gene of isolates from 1999, 2001, and 2002 had a 99.2-99.9% homology, without consistent amino acid variations in the neutralization sites. Phylogenetic analysis of Taiwanese isolates revealed 2 distinct clusters, the 1983-1989 and 1996-2002 isolates. Cross-neutralization tests confirmed the glycoprotein gene sequence analysis results. In conclusion, annual boosters at SNA levels > 32, at more than 2 doses, or at intervals shorter than 6 months are not advisable. The occurrence of frequent small epizootics implies the dominance of BEF virus over host immunity, but not a variant virus.
Journal of Veterinary Medical Science 04/2005; 67(4):411-6. · 0.85 Impact Factor
