1,721,021 research outputs found
Constitutive expression of Atlantic salmon Mx1 protein in CHSE-214 cells confers resistance to Infectious Salmon Anaemia virus
No full textAbstract Infectious salmon anaemia (ISA) is a highly fatal viral disease affecting marine-farmed Atlantic salmon which is caused by ISA virus (ISAV), a fish orthomyxovirus that has recently been assigned to the new genus Isavirus within the family Orthomyxoviridae. Mx proteins are among the interferon (IFN)-induced proteins responsible for the development of an antiviral state in vertebrate cells. We used real-time reverse transcription-polymerase chain reaction (RT-PCR) and Chinook salmon embryo (CHSE-214) cells constitutively expressing Atlantic salmon Mx1 protein (ASMx1) to examine the antiviral properties of ASMx1 against two ISAV strains, NBISA01 and HKS-36, having phenotypically different growth properties (cytopathic vs non-cytopathic) in the CHSE-214 cell line. We present evidence that ISAV is sensitive to ASMx1. CHSE-214 cells constitutively expressing ASMx1 showed increased resistance to infection with the cytopathic ISAV strain NBISA01, manifested as delayed development of cytopathic effects (CPE) and significant reduction in the severity of CPE, as well as a 10-fold reduction in virus yield. However, by real-time RT-PCR we observed no significant difference in the mean threshold cycle (Ct) values of ISAV RNA levels, suggesting that the ASMx1 activity on ISAV occurs at the post-transcription steps of virus replication, possibly in the cytoplasm.</p
Differential detection of infectious bursal disease virus serotypes, using cDNA probes to VP2 coding region
No full textTwo nonoverlapping clones, pOH405 and pOH632, containing cDNA inserts in the VP2 coding region of genome segment A were selected from a cDNA library prepared from the double-stranded RNA genome of the OH strain of infectious bursal disease virus (IBDV) of serotype 2. Clone pOH405, which is located in the hypervariable segment of VP2, is 328 base pairs long, has nucleotide sequence homology of 72 to 73%, and amino acid sequence homology of 64 to 67% with IBDV strains of serotype 1. Clone pOH632, which is located in the highly conserved C-terminal part of VP2, is 230 base pairs long, has nucleotide sequence homology of 87 to 88%, and amino acid sequence homology of 100% with IBDV serotype 1. The lower detection limit of 32P-labeled probes prepared from both clones was 10 ng of OH-IBDV double-stranded RNA, using high-stringency conditions of hybridization (54 C, 50% formamide) and washing (55 C, 0.015M NaCl, 0.0015M trisodium citrate, pH 7.0, with 0.1% sodium dodecyl sulfate), and autoradiography for 24 hours. Under these conditions, the dot-blot hybridization assay for detection of serotype 2 IBDV double-stranded RNA, was 1,000 times more sensitive, using probe pOH632, but only 10 times more sensitive, using probe pOH405, compared with the assay for IBDV serotype 1, using the same probes. Thus, probe pOH632 could differentiate between the 2 IBDV serotypes by nucleic acid hybridization.LR: 20061115; PUBM: Print; JID: 0375011; 0 (DNA Probes); 0 (DNA, Viral); 0 (Viral Structural Proteins); ppublishSource type: Electronic(1
Rapid passage of avian reovirus in one-day-old chicks: clinical and virological findings
No full textTwo avian reoviruses, strain Reo-25 and isolate W3-492 were given by mouth to eight one-day-old chicks. 3-7 days after inoculation, the liver, spleen, pancreas, caecal tonsil and duodenum were collected, weighed and titrated in cell culture for viral content. Tissue homogenates were passaged several times in day-old chicks. Reo-25 virus was passaged at 3-day and W3-492 virus at 3- and 7- or 14-day intervals. For both Reo-25 and W3-492 viruses, pathological effects and virus yields in tissues decreased with continued passages. In direct comparisons of reovirus W3-492 before chick passage (PO) and after four passages at 7-day intervals (P4) using standardised amounts of virus for inoculation of chickens, no major differences in pathological effects were observed. P4 virus could be recovered from duodenal tissue at 28 days and from liver tissue at 14 days. PO virus could be recovered from duodenal tissue at 14 days and from liver tissue at 10 days..RE: 25 ref.; SC: ZA; CA; VE; 0V; 7A; 0ISource type: Electronic(1) http://upei-resolver.asin-risa.ca?sid=SP:CABI&id=pmid:&id=&issn=0307-9457&isbn=&volume=16&issue=2&spage=213&pages=213-225&date=1987&title=Avian%20Pathology&atitle=Rapid%20passage%20of%20avian%20reovirus%20in%20one-day-old%20chicks%3a%20clinical%20and%20virological%20findings.&aulast=Kibenge&pid=%3Cauthor%3EKibenge%2c%20F%20S%20B%3bDhillon%2c%20A%20S%3C%2Fauthor%3E%3CAN%3E19872296778%3C%2FAN%3E%3CDT%3EJournal%20article%3C%2FDT%3
Observations on polymerase chain reaction amplification of infectious bursal disease virus dsRNA
No full textTwo methods for denaturing double stranded (ds)RNA of infectious bursal disease virus for the purpose of reverse transcribing it were compared: Heat denaturation at 65 degrees C in the presence of DMSO and in the absence of DMSO. As part of the analysis, the nature of cDNA in the two preparations was examined by polymerase chain reaction (PCR) amplification, firstly by varying the number of cycles of PCR, and secondly by re-amplification of serial dilutions of the reaction products. The results show that denaturation of dsRNA in the presence of DMSO (method 1) is superior to denaturation without DMSO (method 2) judging by the yield of a specific PCR fragment after 30 cycles, and that the products of method 2 can be re-amplified, albeit poorly, with the generation of heterologous products.LR: 20061115; PUBM: Print; JID: 8005839; 0 (RNA, Double-Stranded); 0 (RNA, Viral); 67-68-5 (Dimethyl Sulfoxide); ppublishSource type: Electronic(1
Sequence conservation in the RNA polymerase gene of infectious bursal disease viruses
No full textEleven of thirteen infectious bursal disease viruses (IBDV) were shown to be similar following polymerase chain reaction (PCR) amplification of two regions (293 bp and 651 bp cDNA fragments) in genomic segment B and restriction enzyme analysis of the PCR products. Two IBDV strains differed from the eleven viruses when examined by BstEII digestion of the 293 bp PCR product, but all thirteen viruses were similar using the 651 bp cDNA fragment and digestion with AvaI. Because segment B encodes replicating enzyme(s), the results suggest that its sequences may be highly conserved among IBDV isolates.LR: 20061115; PUBM: Print; GENBANK/L19502; JID: 7506870; 0 (DNA Primers); 0 (RNA, Double-Stranded); EC 2.7.7.6 (DNA-Directed RNA Polymerases); EC 3.1.21.- (endodeoxyribonuclease EcaI); EC 3.1.21.4 (Deoxyribonucleases, Type II Site-Specific); ppublishSource type: Electronic(1
Restriction fragment profiles of genome segment A of infectious bursal disease virus correlate with serotype and geographical origin of avibirnaviruses
No full textPrevious analysis of the two serotypes of infectious bursal disease virus (IBDV) have demonstrated the correlation between antigenicity and similarities of nucleotide and amino acid sequences of the VP2 coding region in genome segment A. Restriction fragment profiles of genomic segment A cDNA of five IBDV isolates (QC-2 and QT-1 of serotype 1, SK9, and Nos. 39 and 52 of serotype 2) were determined in order to establish the genetic relationship of these viruses to other avibirnaviruses. The restriction fragment profiles using three of seven restriction enzymes (SacI which cuts in the VP2 region, DraIII which cuts in the VP3 region, and EcoRI which cuts in the VP4 region) were used to place QC-2, QT-1, SK9, No. 39, and No. 52 within the phylogenetic tree among seven other avibirnaviruses of known sequence. The two IBDV serotypes corresponded to two genotypes on the basis of the presence or absence of the SacI restriction site. The serotype 1 cluster of strains was further differentiated into five minor clusters on the basis of the PstI, EcoRI, BamHI, HindIII, DraIII, and Bsu361 restriction sites, which emphasized the geographical origins of the strains. It is concluded that restriction analysis of cDNA of the whole viral genomic segment A allows differentiation of IBDV isolates on the basis of their antigenicity and geographical origin.LR: 20061115; PUBM: Print; JID: 0372707; 0 (DNA, Complementary); 0 (RNA, Viral); ppublishSource type: Electronic(1
A comparison of the pathogenicity of four avian reoviruses in chickens
No full textFour avian reoviruses were orally inoculated into 1-day-old chickens to determine pathogenicity, virus persistence in the intestinal tract, and effects on body weight gains. Avian reoviruses Reo-25 and W3-492 belonged to two separate serotypes, and viruses TC 897 and W3-410 were antigenically related to W3-492. Isolate W3-492, which was highly pathogenic, was very rarely recovered from cloacal swabs collected 2 weeks postinoculation, but inoculated chickens gained significantly less weight (P less than or equal to 0.001) than uninoculated controls during the 5-week test study. Isolate Reo-25 persisted the longest in the intestinal tract, and isolates TC 897 and W3-410, of intermediate persistence, had no significant effect on body weights. There was no apparent correlation between serotype and pathotype of avian reoviruses.LR: 20061115; PUBM: Print; JID: 0370617; ppublishSource type: Electronic(1
Reovirus-induced tenosynovitis in chickens: the effect of breed
No full textThe effect of breed of chicken on infection with an arthrotropic avian reovirus (strain R2) was studied by oral or footpad inoculation of day-old chicks of the SPF light-hybrid, commercial White Leghorn egg-layer, and commercial Ross I broiler breeds, observed to 12 weeks of age. Although most inoculated birds of all three breeds developed swelling of one or both legs below the hock joint at 3 to 4 weeks of age, gross lesions of tenosynovitis became progressively more severe and extended above the joints only in broilers, whereas in most orally-infected SPF and commercial light chickens gross lesions were intermittently severe and regressed with time. Cloacal virus shedding continued up to 2 weeks in the lighter breeds and 3 weeks after infection in broilers. From a small proportion of infected chickens, reovirus was also recovered from heart, pancreas and caecal tonsils. In all breeds, the tissue in which virus persisted longest was the hock joint/tendon. There was a poor correlation between isolation of virus and the presence of gross lesions in chickens of 12 weeks of age, especially in broilers. Virus-neutralization tests demonstrated that seroconversion in the lighter breeds occurred mainly at 3 weeks, and in broilers at 4 weeks after infection. In all three breeds the footpad infection resulted in poorer growth than in the control and oral-infection groups. Oral infection had no apparent effect on growth rate. The greater susceptibility of broilers to reovirus infection is discussed..RE: 30 ref.; SC: ZA; CA; VE; 0V; 7A; 0ISource type: Electronic(1) http://upei-resolver.asin-risa.ca?sid=SP:CABI&id=pmid:&id=&issn=0307-9457&isbn=&volume=13&issue=3&spage=511&pages=511-528&date=1984&title=Avian%20Pathology&atitle=Reovirus-induced%20tenosynovitis%20in%20chickens%3a%20the%20effect%20of%20breed.&aulast=Jones&pid=%3Cauthor%3EJones%2c%20R%20C%3bKibenge%2c%20F%20S%20B%3C%2Fauthor%3E%3CAN%3E19842247265%3C%2FAN%3E%3CDT%3EJournal%20article%3C%2FDT%3
The 5'-terminal 32 basepairs conserved between genome segments A and B contain a major promoter element of infectious bursal disease virus
No full textThe regions of the infectious bursal disease virus (IBDV) genome with regulatory function are not known. In the present study, progressively deleted lengths of the 5' noncoding region of segment A were constructed in pGL3 vectors having SV40 enhancer or promoter, and a luciferase (LUC) reporter gene. Transient transfections of the constructs made in a promoter-less pGL3-Enhancer vector when transfected in Vero cells and the lysates assayed for LUC expression, allowed the localization of maximal activity to the 32-nucleotide stretch (precursor polyprotein ORF positions -131 to -100), which is highly conserved at the 5' end of both genome segments. This fragment, when evaluated in parallel in an enhancer-less pGL3-Promoter vector demonstrated no activity. To determine if this region is recognized by IBDV replicative proteins, we engineered modifications in an enhancer-less pGL3-Promoter vector where the terminal 32-bp fragment, the full-length noncoding region, or the noncoding region with the 32-bp fragment deleted was positioned in either the plus-sense or the minus-sense orientation immediately downstream of the SV40 promoter and upstream of the LUC gene. Transfections of these constructs in IBDV-infected and uninfected Vero cells resulted in the endogenous generation of recombinant viral-LUC RNAs containing the 5' terminal viral RNA sequences in either the plus-sense or the minus-sense orientation. LUC assays of the infected cell lysates showed up-regulated expression of LUC only with constructs containing the 32-bp fragment in the minus-sense orientation. Deletion of this 32-bp fragment abolished such LUC expression. We therefore conclude that the 5'-terminal 32 base pairs of genomic segment A contain a major promoter element in IBDV. In addition, our results show that IBDV replicative proteins recognize and transcribe single-stranded RNA in vivo.LR: 20061115; PUBM: Print; JID: 7506870; ppublishSource type: Electronic(1
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