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C A Dangler

Publications and source records attributed to C A Dangler.

49 records · Page 3Linked to original sources
PubMed1994

Spatial and biological characteristics of between-herd transmission of Aujeszky's disease virus.

The eradication of Aujeszky's disease virus (ADV) in Pennsylvania has suffered because the modes of viral transmission between herds are unknown. Our objective was to identify the major risk factors involved in viral transmission by comparing proportions of operation type, density, quarantine level, and vaccination status of neighboring herds for infected case and uninfected control herds. Multivariate analysis demonstrated an elevated risk of infection with local increases in the proportion of finishing floors. A local increase in the proportion of ADV-quarantined herds was shown to be a significant risk factor, as was an increase in the proportion of vaccinating herds. A reduced risk of infection was found with local increases in the number of feeder pig producer operations.

Animal Husbandry↗
PubMed1993

Direct isolation and identification of recombinant pseudorabies virus strains from tissues of experimentally co-infected swine.

Tissue homogenates were obtained from swine co-infected with 2 vaccine strains of pseudorabies virus (PRV). Viral isolates derived by serial plaque purification directly from tissue homogenates, without an intervening step of isolation and amplification on cell cultures, were characterized as recombinant and parental PRV genotypes on the basis of thymidine kinase and glycoprotein X gene combinations. Use of limiting dilutions and recovery of virus isolates as individual plaques minimized the likelihood of in vitro recombination serving as a confounding source of recombinant PRV. The thymidine kinase and glycoprotein X gene sequences were classified as wild-type or deleted, using a battery of polymerase chain reaction assays. Results substantiate the observation that PRV vaccine strains can form genetic recombinants in vivo after experimentally induced co-infection.

Animals↗
PubMed1992

Hybridization relatedness of Israeli and U.S. bluetongue (BLU) serotypes using cDNA probes from BLU virus strain 11-UC8.

Partial cDNA clones representing 47%, 96%, and 98% of genome segments 7, 9, and 10, respectively, of a US bluetongue virus (BLU) 11 virulent strain were used to study, for the first time, the genetic relationships between Israeli BLU proto-serotypes and field isolates, and US BLU proto-serotypes. Their usefulness as group-specific identification probes was also determined. The viral nucleic acid was extracted from the infected cells and the purified dsRNA genome segments were fractionated by polyacrylamide gel electrophoresis, transferred to a nylon membrane and hybridized to the 32P labeled DNA probes. The three probes recognized all the samples tested. Genome segment 7, that code for the mayor inner capsid protein VP7, showed the most variation in the hybridization signal with the US proto-serotypes and all the Israeli samples studied. The genome segments 9 and 10 that code for the minor inner capsid protein VP6 and the nonstructural protein NS3, respectively, were highly conserved in all the samples tested despite their distant geographical regions of origin. The last two mentioned clones showed to be good group-specific probes for the identification of BLU samples from Israel and United States. The obtained cloned genetic probes were also tested against US epizootic haemorrhagic disease virus (EHDV) serotype 1 and 2 viral dsRNA, a distantly related orbivirus. None of them hybridized with the viral dsRNA of these two viruses.

Animals↗
PubMed1992

Genotypic screening of pseudorabies virus strains for thymidine kinase deletions by use of the polymerase chain reaction.

Genetic recombination between field strains and vaccine strains of pseudorabies virus (PRV) has been suggested as a scenario that might arise from use of deletion-mutant modified-live vaccine strains, particularly those strains attenuated by deletions within the thymidine kinase (TK-) gene locus. To address this hypothesis experimentally, it is necessary to screen large numbers of PRV isolates for their TK genotype. Techniques to detect the native TK genotype are routinely used in molecular virology laboratories, but are time-consuming. We adapted the polymerase chain reaction to define the genotypic status of PRV isolates with regard to the presence or absence of deletions in the TK gene locus. Used in tandem with the existing glycoprotein-specific ELISA that discriminate between PRV-vaccinated and field strain-infected swine populations, the described technique may help to clarify whether vaccine-derived recombinants are generated under natural conditions and after normal vaccine usage.

Animals↗
PubMed1992

Comparison of slot blot nucleic acid hybridization, immunofluorescence, and virus isolation techniques to detect bluetongue virus in blood mononuclear cells from cattle with experimentally induced infection.

A slot blot hybridization technique was applied for detection of bluetongue virus (BTV) in blood mononuclear cells (BMNC) obtained from cattle with experimentally induced infection. This technique lacked sensitivity to detect the viral nucleic acid directly in clinical specimens. When aliquots of mononuclear cells from these cattle were cultivated in vitro for 10 days to amplify virus titer, only 33.3% of the samples collected during viremia gave a positive signal in the slot blot hybridization format. By contrast, results for 34.3% of noncultured and 63.3% of cultured mononuclear cell samples collected during viremia were positive by immunofluorescence. The average number of infected cells, as detected by immunofluorescence in the noncultured mononuclear cell samples, was 1 to 5/300,000, and was usually > 10/300,000 in the cultured cell samples. Virus was isolated from all postinoculation blood samples obtained from 4 heifers that were seronegative at the time of inoculation, but was not isolated from any of the preinoculation samples, or from any of the postinoculation samples obtained from 2 heifers that were seropositive at the time of inoculation. When virus isolation was attempted from separated mononuclear cells in 2 heifers, 43.7% of the noncultured and 87.5% of the cultured samples had positive results.

Animals↗
PubMed1990

Identifying bluetongue virus ribonucleic acid sequences by the polymerase chain reaction.

A primer-directed nucleic acid amplification reaction, commonly known as the polymerase chain reaction (PCR), has been adapted to the identification of bluetongue virus (BTV) RNA. The protocol described in this article is designed for the detection of a purported globally-conserved, serogroup specific nucleic acid sequence within the BTV genome. Due to the double-stranded RNA composition of the BTV genome, the original polymerase chain reaction protocol has been modified to include chemical denaturation and reverse transcription steps that allow selective amplification from this unique template molecule. The amplification procedure yields a 210 base pair product when tested on samples of RNA from the prototypic strains of U.S. BTV serotypes 2, 10, 11, 13, 17. The amplification product is tentatively identified by agarose gel electrophoresis of the PCR samples. Final positive identification of the amplified BTV-specific product is determined by Southern blot hybridization of the PCR samples. RNA samples from uninfected cell culture controls and from cell cultures infected with two serotypes of epizootic hemorrhagic disease of deer (EHDV) yielded negative results. Preliminary experiments to quantify the threshold of sensitivity of this protocol, as described here, indicate positive detection of the BTV target RNA sequence at a level of less than 2 fg, or the amount of target sequence derived from less than 7500 viral particles. While this falls short of the theoretical limit of sensitivity of the PCR, the enhanced threshold of sensitivity of the PCR reaction compared to standard nucleic acid hybridization methodology suggests that rapid detection of BTV RNA directly within clinical specimens may be feasible with further refinements. Potential methods of enhancing this reaction are discussed.

Animals↗
PubMed1990

Limitations of in situ hybridization for the detection of bluetongue virus in blood mononuclear cells.

In situ nucleic acid hybridization was tested for the ability to detect bluetongue virus (BTV) nucleic acids in blood mononuclear cells. A standard protocol was devised and applied to the demonstration of BTV genetic sequences in cultured bovine mononuclear cells that had been infected in vitro. In situ hybridization using biotinylated single-stranded RNA probes, in the presence of 50% formamide at 50 C, demonstrated an intense, positive signal in 0.001-0.01% of the BTV-infected cultured mononuclear cells. The protocol was applied to isolated mononuclear cells from an experimentally infected heifer. No infected cells were observed by this method, although the blood specimens were obtained during peak viremia.

Animals↗
PubMed1989

Recombinant DNA probe for serotype-specific identification of bluetongue virus 17.

The double-stranded RNA genome from 117 field isolates of bluetongue virus (BTV) serotypes 10, 11, 13, and 17 was blotted onto nitrocellulose paper and hybridized with a radioactively labeled cloned copy of DNA genome segment 2 of BTV-17. Viral RNA from BTV prototype strains 2, 10, 11, 13, and 17 were used as controls. The probe hybridized only with the viral RNA from prototype BTV-17 virus and field isolates of BTV-17. There was no cross hybridization with field isolates of BTV serotypes 10, 11, and 13. A complementary DNA probe developed from genes coding for BTV serotype specificity was effectively used in a slot-blot hybridization system for efficiently characterizing the viral serotype.

Animals↗
PubMed1988

Rapid identification of bluetongue virus by nucleic acid hybridization in solution.

A liquid phase hybridization format has been adapted for the identification of bluetongue virus (BTV) with respect to serogroup and serotype. In this paper we present experiments which demonstrate the identification of bluetongue virus with respect to both serogroup and serotype. BTV serogroup-specific and BTV serotype 17-specific single-stranded RNA probes labeled with 35S-CTP were used to identify BTV viral nucleic acids in infected cell culture lysates. Cell lysates were prepared for testing by rapid solubilization in a guanidine isothiocyanate/EDTA/dithiothreitol medium. Hybridizations were done in 12.5 microliter reaction volumes for 2 h or overnight using nanogram quantities of probe. An RNAse solution was added subsequent to hybridization and incubated for 30 min. TCA-precipitable counts were collected and quantitated by scintillation counting. Specific hybridization signals were obtained in as little as 3 h in assays consisting of uninfected controls, cells infected with different BTV serotypes, and cells infected with a strain of epizootic hemorrhagic disease of deer (EHDV-1). By using asymmetric single-stranded RNA probes we were able to distinguish viral mRNA and viral double-stranded RNA components. We propose this technique as an ancillary or replacement procedure for the confirmation and classification of viral isolates using an appropriate battery of nucleic acid probes. Potential applications and methods of enhancing the technique are discussed.

Bluetongue virus↗
PubMed1986

Multiple bluetongue virus-cloned genetic probes: application to diagnostics and bluetongue virus genetic relationships.

A shotgun-cloning method incorporating all 10 bluetongue virus genome segments can simultaneously produce complete and partial copies of any of the genome segments. We report here 4 different cloned probes derived from 3 genome segments and individually defined by different hybridization recognition capabilities. One probe hybridized strongly with all 5 United States prototype strains of the 5 different bluetongue virus (BTV) serotypes existing in the United States and, as such, is a strong candidate for a broad BTV diagnostic probe in the United States. Another probe derived from genome segment 2 of BTV-17 hybridized only with the BTV-17 prototypic serotype, thereby demonstrating serospecific hybridization diagnostic potential. The implications for diagnostic and genetic relationship studies on BTV, using various genetic probes, are discussed.

Animals↗
PubMed1984

Protozoal myeloencephalitis in horses in California.

Three cases of equine protozoal myeloencephalitis were diagnosed over a 12-month period in horses that had never left the state of California. These cases suggest that the disease is enzootic in California.

Animals↗