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The IL-10 homologue encoded by cyprinid herpesvirus 3 is essential neither for viral replication in vitro nor for virulence in vivo.

Ouyang P, Rakus K, Boutier M, Reschner A, Leroy B, Ronsmans M, Fournier G, Scohy S, Costes B, Wattiez R, Vanderplasschen A - Vet. Res. (2013)

Bottom Line: Quantification of viral DNA by real time TaqMan PCR (in the gills and the kidney) and analysis of carp cytokine expression (in the spleen) by RT-qPCR at different times post-infection did not revealed any significant difference between the groups of fish infected with the three virus genotypes.Similarly, histological examination of the gills and the kidney of infected fish revealed no significant differences between fish infected with ORF134 deleted virus versus fish infected with the control parental or revertant strains.All together, the results of the present study demonstrate that the IL-10 homologue encoded by CyHV-3 is essential neither for viral replication in vitro nor for virulence in common carp.

View Article: PubMed Central - HTML - PubMed

Affiliation: Immunology-Vaccinology (B43b), Department of Infectious and Parasitic Diseases (B43b), Faculty of Veterinary Medicine, University of Liège, Liège, B-4000, Belgium. a.vdplasschen@ulg.ac.be.

ABSTRACT
Cyprinid herpesvirus 3 (CyHV-3), a member of the family Alloherpesviridae, is the causative agent of a lethal disease in common and koi carp. CyHV-3 ORF134 encodes an interleukin-10 (IL-10) homologue. The present study was devoted to this ORF. Transcriptomic analyses revealed that ORF134 is expressed as a spliced gene belonging to the early-late class. Proteomic analyses of CyHV-3 infected cell supernatant demonstrated that the ORF134 expression product is one of the most abundant proteins of the CyHV-3 secretome. To investigate the role of ORF134 in viral replication in vitro and in virulence in vivo, a deleted strain and a derived revertant strain were produced using BAC cloning technologies. The recombinant ORF134 deleted strain replicated in vitro comparably to the parental and the revertant strains. Infection of fish by immersion in water containing the virus induced comparable CyHV-3 disease for the three virus genotypes tested (wild type, deleted and revertant). Quantification of viral DNA by real time TaqMan PCR (in the gills and the kidney) and analysis of carp cytokine expression (in the spleen) by RT-qPCR at different times post-infection did not revealed any significant difference between the groups of fish infected with the three virus genotypes. Similarly, histological examination of the gills and the kidney of infected fish revealed no significant differences between fish infected with ORF134 deleted virus versus fish infected with the control parental or revertant strains. All together, the results of the present study demonstrate that the IL-10 homologue encoded by CyHV-3 is essential neither for viral replication in vitro nor for virulence in common carp.

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Effect of ORF134 deletion on viral growth in vitro. Replication kinetics of CyHV-3 ORF134 recombinant strains were compared with those of the FL BAC revertant strain using a multi-step growth assay (see Materials and methods). The data presented are the means ± standard errors of triplicate measurements.
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Figure 7: Effect of ORF134 deletion on viral growth in vitro. Replication kinetics of CyHV-3 ORF134 recombinant strains were compared with those of the FL BAC revertant strain using a multi-step growth assay (see Materials and methods). The data presented are the means ± standard errors of triplicate measurements.

Mentions: In order to investigate the putative effects of the recombination processes on viral growth in vitro, the FL BAC revertant, the FL BAC revertant ORF134 Del and the FL BAC revertant ORF134 Rev were compared using a multi-step growth assay (Figure 7). All viruses tested exhibited similar growth curves (P ≤ 0.05), leading to the conclusion that ORF134 deletion does not affect viral growth in vitro (Figure 7). This observation is consistent with what has been reported for other vIL-10s [23,25]. Taken together, these results indicate that ORF134 is not essential for CyHV-3 replication in vitro and suggest that ORF134 exerts its biological functions in vivo.


The IL-10 homologue encoded by cyprinid herpesvirus 3 is essential neither for viral replication in vitro nor for virulence in vivo.

Ouyang P, Rakus K, Boutier M, Reschner A, Leroy B, Ronsmans M, Fournier G, Scohy S, Costes B, Wattiez R, Vanderplasschen A - Vet. Res. (2013)

Effect of ORF134 deletion on viral growth in vitro. Replication kinetics of CyHV-3 ORF134 recombinant strains were compared with those of the FL BAC revertant strain using a multi-step growth assay (see Materials and methods). The data presented are the means ± standard errors of triplicate measurements.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3750702&req=5

Figure 7: Effect of ORF134 deletion on viral growth in vitro. Replication kinetics of CyHV-3 ORF134 recombinant strains were compared with those of the FL BAC revertant strain using a multi-step growth assay (see Materials and methods). The data presented are the means ± standard errors of triplicate measurements.
Mentions: In order to investigate the putative effects of the recombination processes on viral growth in vitro, the FL BAC revertant, the FL BAC revertant ORF134 Del and the FL BAC revertant ORF134 Rev were compared using a multi-step growth assay (Figure 7). All viruses tested exhibited similar growth curves (P ≤ 0.05), leading to the conclusion that ORF134 deletion does not affect viral growth in vitro (Figure 7). This observation is consistent with what has been reported for other vIL-10s [23,25]. Taken together, these results indicate that ORF134 is not essential for CyHV-3 replication in vitro and suggest that ORF134 exerts its biological functions in vivo.

Bottom Line: Quantification of viral DNA by real time TaqMan PCR (in the gills and the kidney) and analysis of carp cytokine expression (in the spleen) by RT-qPCR at different times post-infection did not revealed any significant difference between the groups of fish infected with the three virus genotypes.Similarly, histological examination of the gills and the kidney of infected fish revealed no significant differences between fish infected with ORF134 deleted virus versus fish infected with the control parental or revertant strains.All together, the results of the present study demonstrate that the IL-10 homologue encoded by CyHV-3 is essential neither for viral replication in vitro nor for virulence in common carp.

View Article: PubMed Central - HTML - PubMed

Affiliation: Immunology-Vaccinology (B43b), Department of Infectious and Parasitic Diseases (B43b), Faculty of Veterinary Medicine, University of Liège, Liège, B-4000, Belgium. a.vdplasschen@ulg.ac.be.

ABSTRACT
Cyprinid herpesvirus 3 (CyHV-3), a member of the family Alloherpesviridae, is the causative agent of a lethal disease in common and koi carp. CyHV-3 ORF134 encodes an interleukin-10 (IL-10) homologue. The present study was devoted to this ORF. Transcriptomic analyses revealed that ORF134 is expressed as a spliced gene belonging to the early-late class. Proteomic analyses of CyHV-3 infected cell supernatant demonstrated that the ORF134 expression product is one of the most abundant proteins of the CyHV-3 secretome. To investigate the role of ORF134 in viral replication in vitro and in virulence in vivo, a deleted strain and a derived revertant strain were produced using BAC cloning technologies. The recombinant ORF134 deleted strain replicated in vitro comparably to the parental and the revertant strains. Infection of fish by immersion in water containing the virus induced comparable CyHV-3 disease for the three virus genotypes tested (wild type, deleted and revertant). Quantification of viral DNA by real time TaqMan PCR (in the gills and the kidney) and analysis of carp cytokine expression (in the spleen) by RT-qPCR at different times post-infection did not revealed any significant difference between the groups of fish infected with the three virus genotypes. Similarly, histological examination of the gills and the kidney of infected fish revealed no significant differences between fish infected with ORF134 deleted virus versus fish infected with the control parental or revertant strains. All together, the results of the present study demonstrate that the IL-10 homologue encoded by CyHV-3 is essential neither for viral replication in vitro nor for virulence in common carp.

Show MeSH
Related in: MedlinePlus