scholarly journals Development and characterization of an infectious cDNA clone of the virulent Bucyrus strain of Equine arteritis virus

2007 ◽  
Vol 88 (3) ◽  
pp. 918-924 ◽  
Author(s):  
Udeni B. R. Balasuriya ◽  
Eric J. Snijder ◽  
Hans W. Heidner ◽  
Jianqiang Zhang ◽  
Jessika C. Zevenhoven-Dobbe ◽  
...  
Keyword(s):  
Reverse Genetics ◽  
Severe Disease ◽  
Plasmid Vector ◽  
High Titre ◽  
Equine Arteritis Virus ◽  
Progeny Virus ◽  
Cdna Clones ◽  
Virulence Determinants ◽  
Severity Of The Disease

Strains of Equine arteritis virus (EAV) differ in the severity of the disease that they induce in horses. Infectious cDNA clones are potentially useful for identification of genetic determinants of EAV virulence; to date, two clones have been derived from a cell culture-adapted variant of the original (Bucyrus) isolate of EAV, and it has previously been shown that recombinant virus derived from one of these (rEAV030) is attenuated in horses. A complete cDNA copy of the genome of the virulent Bucyrus strain of EAV has now been assembled into a plasmid vector. In contrast to rEAV030, recombinant progeny virus derived from this clone caused severe disease in horses, characterized by pyrexia, oedema, leukopenia, high-titre viraemia and substantial nasal shedding of virus. The availability of infectious cDNA clones that produce recombinant viruses of different virulence to horses will facilitate characterization of the virulence determinants of EAV through reverse genetics.

scholarly journals De-Coding the Contributions of the Viral RNAs to Alphaviral Pathogenesis

Pathogens ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 771
Author(s):  
Autumn T. LaPointe ◽  
Kevin J. Sokoloski
Keyword(s):  
Host Response ◽  
Severe Disease ◽  
Review Article ◽  
Structural Proteins ◽  
Virulence Determinants ◽  
Healthy Individuals ◽  
Viral Rnas ◽  
Positive Sense ◽  
Identification And Characterization

Alphaviruses are positive-sense RNA arboviruses that are capable of causing severe disease in otherwise healthy individuals. There are many aspects of viral infection that determine pathogenesis and major efforts regarding the identification and characterization of virulence determinants have largely focused on the roles of the nonstructural and structural proteins. Nonetheless, the viral RNAs of the alphaviruses themselves play important roles in regard to virulence and pathogenesis. In particular, many sequences and secondary structures within the viral RNAs play an important part in the development of disease and may be considered important determinants of virulence. In this review article, we summarize the known RNA-based virulence traits and host:RNA interactions that influence alphaviral pathogenesis for each of the viral RNA species produced during infection. Overall, the viral RNAs produced during infection are important contributors to alphaviral pathogenesis and more research is needed to fully understand how each RNA species impacts the host response to infection as well as the development of disease.

scholarly journals De-Coding the Contributions of the Viral RNAs to Alphaviral Pathogenesis

2021 ◽  
Author(s):  
Autumn T. LaPointe ◽  
Kevin J. Sokoloski
Keyword(s):  
Host Response ◽  
Severe Disease ◽  
Review Article ◽  
Structural Proteins ◽  
Virulence Determinants ◽  
Healthy Individuals ◽  
Viral Rnas ◽  
Positive Sense ◽  
Identification And Characterization

Alphaviruses are positive-sense RNA arboviruses that are capable of causing severe disease in otherwise healthy individuals. There are many aspects of viral infection that determine pathogenesis and major efforts regarding the identification and characterization of virulence determinants have largely focused on the roles of the nonstructural and structural proteins. Nonetheless, the viral RNAs of the alphaviruses themselves play important roles in regard to virulence and pathogenesis. In particular, many sequences and secondary structures within the viral RNAs play an important part in the development of disease and may be considered important determinants of virulence. In this review article, we summarize the known RNA-based virulence traits and host:RNA interactions that influence alphaviral pathogenesis for each of the viral RNA species produced during infection. Overall, the viral RNAs produced during infection are important contributors to alphaviral pathogenesis and more research is needed to fully understand how each RNA species impacts the host response to infection as well as the development of disease.

scholarly journals A virulent and pathogenic infectious clone of Senecavirus A

2021 ◽  
Vol 102 (8) ◽  
Author(s):  
Maureen H. V. Fernandes ◽  
Marcelo de Lima ◽  
Lok R. Joshi ◽  
Diego G. Diel
Keyword(s):  
Reverse Genetics ◽  
Infectious Clone ◽  
Disease Onset ◽  
Clinical Signs ◽  
Finishing Pigs ◽  
Virulence Determinants ◽  
Infection Dynamics ◽  
Rna Polymerase Promoter

Senecavirus A (SVA) is a picornavirus that circulates in swine populations worldwide causing vesicular disease (VD) in affected animals. Here we developed a reverse genetics system for SVA based on the well-characterized wild-type SVA strain SD15-26 (wt SVA SD15-26). The full-length cDNA genome of SVA was cloned into a plasmid under a T7 RNA polymerase promoter. Following in vitro transcription, the genomic viral RNA was transfected into BHK-21 cells and rescue of infectious virus (rSVA SD15-26) was shown by inoculation of highly susceptible H1299 cells. In vitro characterization of the rSVA SD15-26 showed similar replication properties and protein expression levels as the wt SVA SD15-26. A pathogenesis study was conducted in 15-week-old finishing pigs to evaluate the pathogenicity and infection dynamics of the rSVA SD15-26 virus in comparison to the wt SVA SD15-26. Animals from both rSVA- and wt SVA SD15-26-inoculated groups presented characteristic SVA clinical signs (lethargy and lameness) followed by the development of vesicular lesions on the snout and/or feet. The clinical outcome of infection, including disease onset, severity and duration was similar in rSVA- and the wt SVA SD15-26-inoculated animals. All animals inoculated with rSVA or with wt SVA SD15-26 presented a short-term viremia, and animals from both groups shed similar amounts of virus in oral and nasal secretion, and faeces. Our data demonstrates that the rSVA SD5-26 clone is fully virulent and pathogenic in pigs, presenting comparable pathogenesis and infection dynamics to the wt SVA SD15-26 strain. The infectious clone generated here is a useful platform to study virulence determinants of SVA, and to dissect other aspects of SVA infection biology, pathogenesis and persistence.

scholarly journals Genetic characterization of equine arteritis virus during persistent infection of stallions

2004 ◽  
Vol 85 (2) ◽  
pp. 379-390 ◽  
Author(s):  
Udeni B. R. Balasuriya ◽  
Jodi F. Hedges ◽  
Victoria L. Smalley ◽  
Andrea Navarrette ◽  
William H. McCollum ◽  
...  
Keyword(s):  
Persistent Infection ◽  
Reproductive Tract ◽  
Genetic Characterization ◽  
Neutralization Assay ◽  
High Titre ◽  
Equine Arteritis Virus ◽  
Replicase Gene ◽  
Defective Interfering Particles ◽  
Genomic Deletions

Equine arteritis virus (EAV) causes a persistent infection of the reproductive tract of carrier stallions. The authors determined the complete genome sequences of viruses (CW96 and CW01) that were present 5 years apart in the semen of a carrier stallion (CW). The CW96 and CW01 viruses respectively had only 85·6 % and 85·7 % nucleotide identity to the published sequence of EAV (EAV030). The CW96 and CW01 viruses had two 1 nt insertions and a single 1 nt deletion in the leader sequence, and a 3 nt coding insertion in ORF1a; thus their genomes included 12 708 nt as compared to the 12 704 nt in EAV030. Variation between viruses present in the semen of stallion CW and EAV030 was especially marked in the replicase gene (ORF1a and 1b), and the greatest variation occurred in the portion of ORF1a encoding the nsp2 protein. The ORFs 3 and 5, which respectively encode the GP3 and GP5 envelope proteins, showed greatest variation amongst ORFs encoding structural EAV proteins. Comparative sequence analyses of CW96 and CW01 indicated that ORFs 1a, 1b and 7 were highly conserved during persistent infection, whereas there was substantial variation in ORFs 3 and 5. Although the variation that occurs in ORF5 results in the emergence of novel phenotypic viral variants as determined by neutralization assay, all variants were neutralized by high-titre polyclonal equine antisera, suggesting that immune evasion is unlikely to be responsible for the establishment of persistent EAV infection of carrier stallions. Northern blot analyses of RNA extracted from cell culture propagated viruses isolated from 10 different persistently infected stallions failed to demonstrate any large genomic deletions, suggesting that defective interfering particles are also unlikely to be important in either the maintenance or clearance of persistent EAV infection of the reproductive tract of carrier stallions.

scholarly journals Identification of a Novel Structural Protein of Arteriviruses

1999 ◽  
Vol 73 (8) ◽  
pp. 6335-6345 ◽  
Author(s):  
Eric J. Snijder ◽  
Hans van Tol ◽  
Ketil W. Pedersen ◽  
Martin J. B. Raamsman ◽  
Antoine A. F. de Vries
Keyword(s):  
Reverse Genetics ◽  
Equine Arteritis Virus ◽  
Progeny Virus ◽  
Replicase Gene ◽  
Structural Protein ◽  
Reading Frame ◽  
Novel Gene ◽  
E Protein ◽  
C Terminus ◽  
Infected Cells

ABSTRACT Arteriviruses are positive-stranded RNA viruses with an efficiently organized, polycistronic genome. A short region between the replicase gene and open reading frame (ORF) 2 of the equine arteritis virus (EAV) genome was previously assumed to be untranslated. However, here we report that this segment of the EAV genome contains the 5′ part of a novel gene (ORF 2a) which is conserved in all arteriviruses. The 3′ part of EAV ORF 2a overlaps with the 5′ part of the former ORF 2 (now renamed ORF 2b), which encodes the GS glycoprotein. Both ORF 2a and ORF 2b appear to be expressed from mRNA 2, which thereby constitutes the first proven example of a bicistronic mRNA in arteriviruses. The 67-amino-acid protein encoded by EAV ORF 2a, which we have provisionally named the envelope (E) protein, is very hydrophobic and has a basic C terminus. An E protein-specific antiserum was raised and used to demonstrate the expression of the novel gene in EAV-infected cells. The EAV E protein proved to be very stable, did not form disulfide-linked oligomers, and was not N-glycosylated. Immunofluorescence and immunoelectron microscopy studies showed that the E protein associates with intracellular membranes both in EAV-infected cells and upon independent expression. An analysis of purified EAV particles revealed that the E protein is a structural protein. By using reverse genetics, we demonstrated that both the EAV E and GS proteins are essential for the production of infectious progeny virus.

scholarly journals Isolation and characterization of cDNA clones corresponding to two different human apoC-III alleles.

1985 ◽  
Vol 26 (4) ◽  
pp. 451-456
Author(s):  
S K Karathanasis ◽  
V I Zannis ◽  
J L Breslow
Keyword(s):  
Cdna Clones ◽  
Isolation And Characterization

scholarly journals Rabbit C-reactive protein. Biosynthesis and characterization of cDNA clones.

1986 ◽  
Vol 261 (12) ◽  
pp. 5473-5479 ◽  
Author(s):  
C Syin ◽  
E C Gotschlich ◽  
T Y Liu
Keyword(s):  
Protein Biosynthesis ◽  
Cdna Clones ◽  
C Reactive Protein ◽  
Reactive Protein

scholarly journals Molecular cloning and characterization of human atrial and ventricular myosin alkali light chain cDNA clones.

1988 ◽  
Vol 263 (27) ◽  
pp. 13930-13936
Author(s):  
M Kurabayashi ◽  
I Komuro ◽  
H Tsuchimochi ◽  
F Takaku ◽  
Y Yazaki
Keyword(s):  
Molecular Cloning ◽  
Light Chain ◽  
Cdna Clones ◽  
Ventricular Myosin
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