Macromolecular synthesis in bluetongue virus infected cells. I. Virus-specific ribonucleic acid synthesis

Both virus-specific double-stranded and single-stranded ribonucleic acid (RNA) are synthesized during infection. The single-stranded RNA is formed in a large excess of double-stranded RNA and the rate of synthesis is maximal between 10 and 13 hours after infection. The single-stranded RNA is associated with the polyribosomes and consists of components with sedimentation constants varying between 12S and 22S. Hybridization of single-stranded RNA with double-stranded RNA indicated that the single-stranded RNA is probably messenger RNA. The secondary structure of the double-stranded RNA was verified by optical rotatory dispersion.The journals have been scanned in colour with a HP 5590 scanner; 600 dpi. Adobe Acrobat v.11 was used to OCR the text and also for the merging and conversion to the final presentation PDF-format..mn201

Macromolecular synthesis in bluetongue virus infected cells. II. Host cell metabolism

Infection of L-cells with bluetongue virus results in inhibition of protein and deoxyribonucleic acid synthesis shortly after infection. No inhibition of ribonucleic acid synthesis is observed before 7 hours after infection. The length of the lag phase before the initiation of the inhibition of protein synthesis is dependent upon the number of infecting virus particles. An increase in the multiplicity of infection results in a decrease in the length of the lag phase. No new macromolecular synthesis is required for the induction of inhibition. Inhibition of viral replication by interferon or UV inactivation does not prevent the induction of inhibition. Virus neutralized by antiserum or inactivated by heat or acid treatment is unable to induce the changes in host cell metabolism.The journals have been scanned in colour with a HP 5590 scanner; 600 dpi. Adobe Acrobat v.11 was used to OCR the text and also for the merging and conversion to the final presentation PDF-format..mn201

Bluetongue virus-induced interferon synthesis

Bluetongue virus was found to induce interferon in mouse embryo (ME) cells and in mice. Different strains of bluetongue virus differed in their ability to induce interferon. Interferon production in ME cells commences after a 5 hour lag phase and the cells continue to produce interferon for 20 hours. Isolated double-stranded bluetongue virus RNA was found to induce maximum titres of interferon in mice approximately 4 hours earlier than was the case with whole virus.The articles have been scanned in colour with a HP Scanjet 5590; 300dpi. Adobe Acrobat XI Pro was used to OCR the text and also for the merging and conversion to the final presentation PDF-format

The use of recombinant DNA technology for the development of a bluetongue virus subunit vaccine

The double-stranded RNA gene coding for the surface antigen responsible for inducing neutralising antibodies has been isolated, converted to DNA, and cloned in the plasmid pBR322. So far, only plasmids containing inserts smaller than the gene have been obtained. Possible strategies for the development of a bluetongue virus subunit vaccine are discussed.The articles have been scanned in colour with a HP Scanjet 5590; 600dpi. Adobe Acrobat XI Pro was used to OCR the text and also for the merging and conversion to the final presentation PDF-format

A comparison of an Australian bluetongue virus isolate (CSIRO 19) with other bluetongue virus serotypes by cross-hybridization and cross-immune precipitation

No major differences in size were observed when both the double-stranded RNA and the polypeptides of the Australian bluetongue virus (BTV) isolate CSIRO 19 (BTV-20) were compared with those of other BTV serotypes such as BTV-10 and BTV-4. Minor capsid polypeptide P6 of both BTV-20 and BTV-4, which electrophoreses as a single band on continuous phosphate buffered gels, is separated into 2 distinct bands on discontinuous glycine-buffered gels. This was not the case with BTV-10. Cross-immune precipitation of BTV-20 with BTV-10, BTV-17, BTV-4 and BTV-3 indicated strong immunological cross-reaction of the group-specific antigen P7 of the different serotypes. There was also some cross-immune precipitation of the serotype-specific polypeptide P2 of BTV-20 and BTV-4. This result is in agreement with the observed cross neutralization of these 2 viruses. The main distinction between BTV-20 and the other BTV serotypes was observed in crosshybridization experiments. The homology between the nucleic acid of BTV-20 and other BTV serotypes was less than 30%, whereas homology normally found between BTV serotypes is at least 70%. The hybridization products of the different BTV serotypes were analysed by electrophoresis and fluorography. Two main hybrid segments were observed in all heterologous hybridizations with BTV-20 as compared with 7 hybrid segments in hybridizations between BTV-4 and BTV-10. In order to determine from which genome segment of BTV -20 these 2 hybrid segments were derived, the hybridizations were carried out with individually purified double-stranded RNA segments. These results indicate that the 2 segments of BTV-20 that show the largest homology to corresponding segments of a heterologous BTV serotype are No. 7 and 10.The articles have been scanned in colour with a HP Scanjet 5590; 600dpi. Adobe Acrobat XI Pro was used to OCR the text and also for the merging and conversion to the final presentation PDF-format

Identification of the serotype-specific and group-specific antigens of bluetongue virus

The bluetongue virus (BTV) core particle contains 2 major polypeptides, P3 and P7, and is surrounded by an outer capsid layer that is composed of the 2 major polypeptides, P2 and P5. Analysis of the immune precipitates from soluble ¹⁴C-labelled BTV polypeptides and hyper-immune rabbit and guinea-pig sera indicated that polypeptide P2 precipitates only with homologous BTV sera. This would indicate that P2 is the main determinant of serotype specificity. It was also found that in sheep infected with BTV the P2-precipitating antibodies in the serum correlate with the neutralizing antibody titres, whereas the appearance and subsequent decline of P7-precipitating antibodies correspond well with those of the complement fixing antibodies. This suggests that BTV group specificity, as measured by a complement fixation test, is determined by the core protein P7. This result was supported by the observation that mouse ascitic fluid, which contains a high titre of BTV specific complement fixing antibodies and a very low titre of neutralizing antibodies, contains almost exclusively antibodies that precipitate P7.The articles have been scanned in colour with a HP Scanjet 5590; 600dpi. Adobe Acrobat XI Pro was used to OCR the text and also for the merging and conversion to the final presentation PDF-format

Molecular hybridization studies on the relationships between different serotypes of bluetongue virus and on the difference between the virulent and attenuated strains of the same serotype

Isolates of ³H-labelled messenger RNA of a number of different bluetongue virus serotypes were hybridized with saturating amounts of denatured ³²P-labelled double-stranded RNA of different serotypes. These cross-hybridization products were then analysed by polyacrylamide gel electrophoresis. The results indicate relatively large differences between the various serotypes. Only a few of the genome segments in the different serotypes were completely homologous. Each of the cross- hybridization patterns obtained using the genome of Serotype 10 and any one of the other serotypes was unique and characteristic for the strain under investigation. The patterns furthermore clearly indicated different degrees of homology between the genomes of the different serotypes. The immunological specificity of the serotypes appears to be determined mainly by the second genome segment of the virus while genome segment six could be of secondary importance. These results were supported by a study of the cross-hybridization patterns between different isolates of Serotype 4. Cross-hybridization experiments between virulent and attenuated strains of the same serotype also indicated small differences. In all the serotypes investigated the process of attenuation involved changes in genome segments two and six. This result would tend to implicate the same genome segments in the determination of both the immunological specificity and the virulence of the virus.The articles have been scanned in colour with a HP Scanjet 5590; 300dpi. Adobe Acrobat XI Pro was used to OCR the text and also for the merging and conversion to the final presentation PDF-format

Interferon induction by bluetongue virus and bluetongue virus ribonucleic acid

The stimulation of interferon synthesis by bluetongue virus and by bluetongue virus ribonucleic acid was investigated in order to determine if there is a difference in the mechanism of induction. The molecular mass of the interferon formed after the two induction processes was determined using Sephadex gel filtration. A value of 24 000 was found in both cases. These results suggest that the two induction processes are basically similar and that double-stranded ribonucleic acid is the active inducing principle in both stimulating processes.The articles have been scanned in colour with a HP Scanjet 5590;300dpi. adobe Acrobat XI Pro was used to OCR the text and also for the merging and conversion to the final presentation PDF-format

Studies on the in vitro and the in vivo transcription of the bluetongue virus genome

Bluetongue virus particles, converted to a high density form by the selective removal of two polypeptides from their protein capsids, possess RNA dependent RNA polymerase activity. The enzyme, which can be assayed by its ability to incorporate nucleoside triphosphates into RNA in an in vitro system, is dependent on magnesium ions, is stimulated by the presence of manganese ions and shows maximal activity at 28°C. The product of the in vitro reaction was isolated and shown to consist of ten single-stranded RNA segments which can be hybridized with double-stranded RNA isolated from purified bluetongue virus (BTV). The hybridization product, when analyzed by means of polyacrylamide gel electrophoresis, is indistinguishable from a hybrid obtained using BTV messenger RNA isolated from infected cells. It is therefore deduced that the BTV genome is fully transcribed both in vitro and in vivo by an enzyme present in the viral capsid.The articles have been scanned in colour with a HP Scanjet 5590;300dpi. adobe Acrobat XI Pro was used to OCR the text and also for the merging and conversion to the final presentation PDF-format

The effect of temperature on the in vitro transcriptase reaction of bluetongue virus, epizootic haemorrhagic disease virus and African horsesickness virus

Virions of bluetongue virus (BTV), epizootic haemorrhagic disease virus (EHDV) and African horsesickness virus (AHSV) can be converted to core particles by treatment with chymotrypsin and magnesium. The conversion is characterized by the removal of the 2 outer capsid polypeptides of the virion. The loss of these 2 proteins results in an increase in density from 1,36g/ml to 1,40g/ml on CsC1 gradients. The BTV, EHDV and AHSV core particles have an associated double-stranded RNA dependent RNA transcriptase that appears to transcribe mRNA optimally at 28⁰C. It was found, at least in the case of BTV, that this low temperature preference is not an intrinsic characteristic of the transcriptase, but is due to a temperature-dependent inhibition of transcription at high core concentrations.The articles have been scanned in colour with a HP Scanjet 5590; 600dpi. Adobe Acrobat XI Pro was used to OCR the text and also for the merging and conversion to the final presentation PDF-format