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Cells Jan 2021Avian reovirus p17 (ARV p17) is a non-structural protein known to activate autophagy, interfere with gene transcription and induce a significant tumor cell growth...
Avian reovirus p17 (ARV p17) is a non-structural protein known to activate autophagy, interfere with gene transcription and induce a significant tumor cell growth inhibition in vitro and in vivo. In this study, we show that ARV p17 is capable of exerting potent antiangiogenic properties. The viral protein significantly inhibited the physiological angiogenesis of human endothelial cells (ECs) by affecting migration, capillary-like structure and new vessel formation. ARV p17 was not only able to suppress the EC physiological angiogenesis but also rendered ECs insensitive to two different potent proangiogenic inducers, such as VEGF-A and FGF-2 in the three-dimensional (3D) Matrigel and spheroid assay. ARV p17 was found to exert its antiangiogenic activity by upregulating transcription and release of the well-known tumor suppressor molecule dipeptidyl peptidase 4 (DPP4). The ability of ARV p17 to impact on angiogenesis is completely new and highlights the "two compartments" activity of the viral protein that is expected to hamper the tumor parenchymal/stromal crosstalk. The complex antitumor activities of ARV p17 open the way to a new promising field of research aimed to develop new therapeutic approaches for treating tumor and cancer metastasis.
Topics: Angiogenesis Inhibitors; Animals; Cell Movement; Chickens; Dipeptidyl Peptidase 4; Fibroblast Growth Factor 2; Human Umbilical Vein Endothelial Cells; Humans; Microvessels; Morphogenesis; Neovascularization, Pathologic; Orthoreovirus, Avian; Recombinant Proteins; Transduction, Genetic; Up-Regulation; Viral Nonstructural Proteins
PubMed: 33525607
DOI: 10.3390/cells10020259 -
Frontiers in Veterinary Science 2023The novel duck reovirus (NDRV) emerged in southeast China in 2005. The virus causes severe liver and spleen hemorrhage and necrosis in various duck species, bringing...
The novel duck reovirus (NDRV) emerged in southeast China in 2005. The virus causes severe liver and spleen hemorrhage and necrosis in various duck species, bringing serious harm to waterfowl farming. In this study, three strains of NDRV designated as NDRV-ZSS-FJ20, NDRV-LRS-GD20, and NDRV-FJ19 were isolated from diseased Muscovy ducks in Guangdong and Fujian provinces. Pairwise sequence comparisons revealed that the three strains were closely related to NDRV, with nucleotide sequence identities for 10 genomic fragments ranging between 84.8 and 99.8%. In contrast, the nucleotide sequences of the three strains were only 38.9-80.9% similar to the chicken-origin reovirus and only 37.6-98.9% similar to the classical waterfowl-origin reovirus. Similarly, phylogenetic analysis revealed that the three strains clustered together with NDRV and were significantly different from classical waterfowl-origin reovirus and chicken-origin reovirus. In addition, the analyses showed that the L1 segment of the NDRV-FJ19 strain was a recombinant of 03G and J18 strains. Experimental reproduction of the disease showed that the NDRV-FJ19 strain was pathogenic to both ducks and chickens and could lead to symptoms of hemorrhage and necrosis in the liver and spleen. This was somewhat different from previous reports that NDRV is less pathogenic to chickens. In conclusion, we speculated that the NDRV-FJ19 causing duck liver and spleen necrosis is a new variant of a duck orthoreovirus that is significantly different in pathogenicity from any previously reported waterfowl-origin orthoreovirus.
PubMed: 36998638
DOI: 10.3389/fvets.2023.1124999 -
Vaccines Mar 2021Heart and skeletal muscle inflammation (HSMI), caused by infection with (PRV-1), is a common disease in farmed Atlantic salmon (). Both an inactivated whole virus...
Heart and skeletal muscle inflammation (HSMI), caused by infection with (PRV-1), is a common disease in farmed Atlantic salmon (). Both an inactivated whole virus vaccine and a DNA vaccine have previously been tested experimentally against HSMI and demonstrated to give partial but not full protection. To understand the mechanisms involved in protection against HSMI and evaluate the potential of live attenuated vaccine strategies, we set up a cross-protection experiment using PRV genotypes not associated with disease development in Atlantic salmon. The three known genotypes of PRV differ in their preference of salmonid host species. The main target species for PRV-1 is Atlantic salmon. Coho salmon () is the target species for PRV-2, where the infection may induce erythrocytic inclusion body syndrome (EIBS). PRV-3 is associated with heart pathology and anemia in rainbow trout, but brown trout () is the likely natural main host species. Here, we tested if primary infection with PRV-2 or PRV-3 in Atlantic salmon could induce protection against secondary PRV-1 infection, in comparison with an adjuvanted, inactivated PRV-1 vaccine. Viral kinetics, production of cross-reactive antibodies, and protection against HSMI were studied. PRV-3, and to a low extent PRV-2, induced antibodies cross-reacting with the PRV-1 σ1 protein, whereas no specific antibodies were detected after vaccination with inactivated PRV-1. Ten weeks after immunization, the fish were challenged through cohabitation with PRV-1-infected shedder fish. A primary PRV-3 infection completely blocked PRV-1 infection, while PRV-2 only reduced PRV-1 infection levels and the severity of HSMI pathology in a few individuals. This study indicates that infection with non-pathogenic, replicating PRV could be a future strategy to protect farmed salmon from HSMI.
PubMed: 33800725
DOI: 10.3390/vaccines9030230 -
Veterinaria Italiana May 2021Common pathogens of intensive poultry farms, either parasitic or bacterial, such as Coccidiaor Salmonella, are well known and strictly controlled by veterinary...
Common pathogens of intensive poultry farms, either parasitic or bacterial, such as Coccidiaor Salmonella, are well known and strictly controlled by veterinary management. This case study reports an unusual case of runting stunting syndrome (RSS) observed on a Sicilian poultry farm of broiler chickens during 2019. The investigation was carried out on five chickens which present delayed in body weight and growth performance. Animals showed also difficulty in deambulation and diarrhea. At necropsy, intestinal lesions were detected in three of the five clinical cases. Gut samples were collected and analyzed to identify potential pathogens responsible for the RSS. Presence of viruses was detected by using quantitative reverse transcription PCR (RT‑qPCR), while selected tissues were fixed and embedded in paraffin wax according to routine procedures. All histological sections were stained with hematoxylin‑eosin. RT‑qPCR successfully detected both Chicken astrovirus (CAstV) and Avian orthoreovirus (ARV). Histology evidenced severe specific lesions on the intestinal mucosa in liver and kidneys. Chicken astrovirus and Avian orthoreovirus RNA was also detected in cecal tonsils, kidney and liver, thus implying their possible primary role in inducing the disease. Further studies are needed to evaluate the role of other possible factors (low biosecurity measures, e.g.) and, most of all, the consequences in terms of economic losses and animal health impairment.
Topics: Animals; Astroviridae Infections; Avastrovirus; Chickens; Coinfection; Diagnosis, Differential; Orthoreovirus, Avian; Polymerase Chain Reaction; Poultry Diseases; RNA, Viral; Reoviridae Infections; Sicily
PubMed: 34313103
DOI: 10.12834/VetIt.2222.13654.1 -
Scientific Reports Dec 2020Birds, notably wild ducks, are reservoirs of pathogenic and zoonotic viruses such as influenza viruses and coronaviruses. In the current study, we used metagenomics to...
Birds, notably wild ducks, are reservoirs of pathogenic and zoonotic viruses such as influenza viruses and coronaviruses. In the current study, we used metagenomics to detect and characterise avian DNA and RNA viruses from wild Pacific black ducks, Chestnut teals and Grey teals collected at different time points from a single location. We characterised a likely new species of duck aviadenovirus and a novel duck gyrovirus. We also report what, to the best of our knowledge, is the first finding of an avian orthoreovirus from Pacific black ducks and a rotavirus F from Chestnut teals. Other viruses characterised from the samples from these wild ducks belong to the virus families Astroviridae, Caliciviridae and Coronaviridae. Some of the viruses may have potential cross-species transmissibility, while others indicated a wide genetic diversity of duck viruses within a genus. The study also showed evidence of potential transmission of viruses along the East Asian-Australasian Flyway; potentially facilitated by migrating shorebirds. The detection and characterisation of several avian viruses not previously described, and causing asymptomatic but potentially also symptomatic infections suggest the need for more virus surveillance studies for pathogenic and potential zoonotic viruses in wildlife reservoirs.
Topics: Animals; Animals, Wild; Birds; Ducks; Gyrovirus; Influenza A virus; Influenza in Birds; Metagenome; Metagenomics; Phylogeny
PubMed: 33335272
DOI: 10.1038/s41598-020-79413-9 -
Journal of Virology Jul 2022Mammalian orthoreovirus (reovirus) is a double-stranded RNA (dsRNA) virus which encapsidates its 10 genome segments within a double-layered viral particle. Reovirus...
Mammalian orthoreovirus (reovirus) is a double-stranded RNA (dsRNA) virus which encapsidates its 10 genome segments within a double-layered viral particle. Reovirus infection triggers an antiviral response in host cells which serves to limit viral replication. This antiviral response is initiated by recognition of the incoming viral genome by host sensors present in the cytoplasm. However, how host sensors gain access to the reovirus genome is unclear, as this dsRNA is protected by the viral particle proteins throughout infection. To initiate infection, reovirus particles are endocytosed and the outer viral particle layer is disassembled through the action of host proteases. This disassembly event is required for viral escape into the cytoplasm to begin replication. We show that endosomal proteases are required even late in infection, when disassembly is complete, to induce an immune response to reovirus. Additionally, counter to dogma, our data demonstrate that at least some viral dsRNA genome is exposed and detectable during entry. We hypothesize that some proportion of reovirus particles remain trapped within endosomes, allowing for the breakdown of these particles and release of their genome. We show that rapidly uncoating mutants escape the endosome more rapidly and induce a diminished immune response. Further, we show that particles entering through dynamin-independent pathways evade detection by host sensors. Overall, our data provide new insight into how genomes from entering reovirus particles are detected by host cells. Viruses must infect host cells to replicate, often killing the host cell in the process. However, hosts can activate defenses to limit viral replication and protect the organism. To trigger these host defenses to viral infections, host cells must first recognize that they are infected. Mammalian orthoreovirus (reovirus) is a model system used to study host-virus interactions. This study identifies aspects of host and virus biology which determine the capacity of host cells to detect infection. Notably, entry of reovirus into host cells plays a critical role in determining the magnitude of immune response triggered during infection. Mutants of reovirus which can enter cells more rapidly are better at avoiding detection by the host. Additionally, reovirus can enter cells through multiple routes. Entry through some of these routes also helps reovirus evade detection.
Topics: Animals; Antiviral Restriction Factors; Cell Line; Immunity, Innate; Orthoreovirus, Mammalian; Peptide Hydrolases; RNA, Double-Stranded; Reoviridae; Reoviridae Infections; Viral Proteins; Virus Replication
PubMed: 35867576
DOI: 10.1128/jvi.00917-22 -
Veterinary Research Oct 2021Piscine orthoreovirus-1 (PRV-1) is the causative agent of heart and skeletal muscle inflammation (HSMI) in farmed Atlantic salmon (Salmo salar). However, it has been...
Piscine orthoreovirus-1 (PRV-1) is the causative agent of heart and skeletal muscle inflammation (HSMI) in farmed Atlantic salmon (Salmo salar). However, it has been shown that PRV-1 variants differ in their ability to induce HSMI. The objective of this work was to identify the PRV-1 variants in Norwegian aquaculture and their geographical distribution. Sequencing and subsequent analysis of the five genomic segments (S1, S4, M2, L1 and L2) putatively linked to virulence, made out the basis of the study. Thirty-seven Norwegian PRV-1 isolates were sequenced, and they grouped into eight genogroups based on combinations of the five analyzed genomic segments. Two groups were defined as high-virulent and two low-virulent, based on comparison with PRV-1 reference isolates with known virulence. The remaining four groups were of unknown virulence. The geographic distribution indicated a higher frequency of the high-virulent isolates in the mid- and northern regions. The present study confirms circulation of both high- and low-virulent isolates of PRV-1 in farmed Atlantic salmon in Norway. To reduce the impact of PRV-1 related disease, detection and differentiation between high- and low-virulent genogroups of PRV-1 could be a targeted approach for reduction of high-virulent variants.
Topics: Animals; Aquaculture; Fish Diseases; Genotype; Norway; Orthoreovirus; Reoviridae Infections; Salmo salar; Virulence
PubMed: 34649601
DOI: 10.1186/s13567-021-01000-1 -
Virus Evolution Jan 2021[This corrects the article DOI: 10.1093/ve/veaa054.].
[This corrects the article DOI: 10.1093/ve/veaa054.].
PubMed: 34168895
DOI: 10.1093/ve/veab008 -
Journal of Virology Mar 2022In this work we have determined that heat shock protein 90 (Hsp90) is essential for avian reovirus (ARV) replication by chaperoning the ARV p17 protein. p17 modulates...
p17-Modulated Hsp90/Cdc37 Complex Governs Oncolytic Avian Reovirus Replication by Chaperoning p17, Which Promotes Viral Protein Synthesis and Accumulation of Viral Proteins σC and σA in Viral Factories.
In this work we have determined that heat shock protein 90 (Hsp90) is essential for avian reovirus (ARV) replication by chaperoning the ARV p17 protein. p17 modulates the formation of the Hsp90/Cdc37 complex by phosphorylation of Cdc37, and this chaperone machinery protects p17 from ubiquitin-proteasome degradation. Inhibition of the Hsp90/Cdc37 complex by inhibitors (17-N-allylamino-17-demethoxygeldanamycin 17-AGG, and celastrol) or short hairpin RNAs (shRNAs) significantly reduced expression levels of viral proteins and virus yield, suggesting that the Hsp90/Cdc37 chaperone complex functions in virus replication. The expression levels of p17 were decreased at the examined time points (2 to 7 h and 7 to 16 h) in 17-AAG-treated cells in a dose-dependent manner while the expression levels of viral proteins σA, σC, and σNS were decreased at the examined time point (7 to 16 h). Interestingly, the expression levels of σC, σA, and σNS proteins increased along with coexpression of p17 protein. p17 together with the Hsp90/Cdc37 complex does not increase viral genome replication but enhances viral protein stability, maturation, and virus production. Virus factories of ARV are composed of nonstructural proteins σNS and μNS. We found that the Hsp90/Cdc37 chaperone complex plays an important role in accumulation of the outer-capsid protein σC, inner core protein σA, and nonstructural protein σNS of ARV in viral factories. Depletion of Hsp90 inhibited σA, σC, and p17 proteins colocalized with σNS in viral factories. This study provides novel insights into p17-modulated formation of the Hsp90/Cdc37 chaperone complex governing virus replication via stabilization and maturation of viral proteins and accumulation of viral proteins in viral factories for virus assembly. Molecular mechanisms that control stabilization of ARV proteins and the intermolecular interactions among inclusion components remain largely unknown. Here, we show that the ARV p17 is an Hsp90 client protein. The Hsp90/Cdc37 chaperone complex is essential for ARV replication by protecting p17 chaperone from ubiquitin-proteasome degradation. p17 modulates the formation of Hsp90/Cdc37 complex by phosphorylation of Cdc37, and this chaperone machinery protects p17 from ubiquitin-proteasome degradation, suggesting a feedback loop between p17 and the Hsp90/Cdc37 chaperone complex. p17 together with the Hsp90/Cdc37 complex does not increase viral genome replication but enhances viral protein stability and virus production. Depletion of Hsp90 prevented viral proteins σA, σC, and p17 from colocalizing with σNS in viral factories. Our findings elucidate that the Hsp90/Cdc37 complex chaperones p17, which, in turn, promotes the synthesis of viral proteins σA, σC, and σNS and facilitates accumulation of the outer-capsid protein σC and inner core protein σA in viral factories for virus assembly.
Topics: Animals; Capsid Proteins; Cell Cycle Proteins; Chaperonins; Genome, Viral; HSP90 Heat-Shock Proteins; Orthoreovirus, Avian; Proteasome Endopeptidase Complex; Ubiquitin; Viral Envelope Proteins; Viral Proteins; Virus Replication
PubMed: 35107368
DOI: 10.1128/jvi.00074-22 -
MBio Aug 2021The function of the mammalian orthoreovirus (reovirus) σNS nonstructural protein is enigmatic. σNS is an RNA-binding protein that forms oligomers and enhances the...
The function of the mammalian orthoreovirus (reovirus) σNS nonstructural protein is enigmatic. σNS is an RNA-binding protein that forms oligomers and enhances the stability of bound RNAs, but the mechanisms by which it contributes to reovirus replication are unknown. To determine the function of σNS-RNA binding in reovirus replication, we engineered σNS mutants deficient in RNA-binding capacity. We found that alanine substitutions of positively charged residues in a predicted RNA-binding domain decrease RNA-dependent oligomerization. To define steps in reovirus replication facilitated by the RNA-binding property of σNS, we established a complementation system in which wild-type or mutant forms of σNS could be tested for the capacity to overcome inhibition of σNS expression. Mutations in σNS that disrupt RNA binding also diminish viral replication and σNS distribution to viral factories. Moreover, viral mRNAs only incorporate into viral factories or factory-like structures (formed following expression of nonstructural protein μNS) when σNS is present and capable of binding RNA. Collectively, these findings indicate that σNS requires positively charged residues in a putative RNA-binding domain to recruit viral mRNAs to sites of viral replication and establish a function for σNS in reovirus replication. Viral replication requires the formation of neoorganelles in infected cells to concentrate essential viral and host components. However, for many viruses, it is unclear how these components coalesce into neoorganelles to form factories for viral replication. We discovered that two mammalian reovirus nonstructural proteins act in concert to form functioning viral factories. Reovirus μNS proteins assemble into exclusive factory scaffolds that require reovirus σNS proteins for efficient viral mRNA incorporation. Our results demonstrate a role for σNS in RNA recruitment to reovirus factories and, more broadly, show how a cytoplasmic non-membrane-enclosed factory is formed by an RNA virus. Understanding the mechanisms of viral factory formation will help identify new targets for antiviral therapeutics that disrupt assembly of these structures and inform the use of nonpathogenic viruses for biotechnological applications.
Topics: HEK293 Cells; Humans; Mutation; Organelles; RNA, Viral; RNA-Binding Proteins; Reoviridae; Viral Nonstructural Proteins; Virus Replication
PubMed: 34225484
DOI: 10.1128/mBio.01408-21