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Journal of Fish Diseases Jun 2024The relationship of histopathological changes and the infection of Piscine orthoreovirus 2 (PRV-2) was investigated in coho salmon that were suffering from the...
Clinical symptoms and histopathological changes in coho salmon affected by the erythrocytic inclusion body syndrome (EIBS) are caused by the infection of piscine orthoreovirus 2 (PRV-2).
The relationship of histopathological changes and the infection of Piscine orthoreovirus 2 (PRV-2) was investigated in coho salmon that were suffering from the erythrocytic inclusion body syndrome (EIBS). Immunohistochemical observations revealed abundant σ1 protein of PRV-2 in the spongy layer of the ventricle of the heart, where severe myocarditis was observed. In the spleen, the virus protein was detected in many erythrocytes, some of which were spherical-shaped and apparently dead. The number of erythrocytes was decreased in the spleen compared to the apparently healthy fish. The virus protein was also detected in some erythrocytes in blood vessels. The viral protein was often detected in many macrophages ingesting erythrocytes or dead cell debris in the spleen or in the kidney sinusoids. Large amounts of the viral genomic segment L2 were also detected in these organs by RT-qPCR. Many necrotic foci were found in the liver, although the virus protein was not detected in the hepatocytes. These results suggest that the primary targets of PRV-2 are myocardial cells and erythrocytes and that clinical symptoms such as anaemia or jaundice and histopathological changes such as myocarditis in EIBS-affected coho salmon are caused by PRV-2 infection.
Topics: Animals; Fish Diseases; Reoviridae Infections; Orthoreovirus; Oncorhynchus kisutch; Erythrocytes; Spleen
PubMed: 38481093
DOI: 10.1111/jfd.13939 -
Research in Veterinary Science May 2024In the pork production chain, the control at slaughterhouse aims to ensure safe food thanks to proper hygienic conditions during all steps of the slaughtering....
In the pork production chain, the control at slaughterhouse aims to ensure safe food thanks to proper hygienic conditions during all steps of the slaughtering. Salmonella is one of the main foodborne pathogens in the EU causing a great number of human cases, and pigs also contribute to its spreading. Pig is the main reservoir of the zoonotic hepatitis E virus (HEV) that can be present in liver, bile, feces and even rarely in blood and muscle. The aim of this study was to assess the presence of both Salmonella and HEV in several points of the slaughtering chain, including pig trucks. Other viruses hosted in the gut flora of pigs and shed in feces were also assayed (porcine adenovirus PAdV, rotavirus, norovirus, and mammalian orthoreovirus MRV). Torque teno sus virus (TTSuV) present in both feces, liver and blood was also considered. Four Italian pig abattoirs were sampled in 12 critical points, 5 of which were the outer surface of carcasses before processing. HEV and rotavirus (RVA) were not detected. Norovirus was detected once. Salmonella was detected in two of the 4 abattoirs: in the two lairage pens, in the site of evisceration and on one carcass, indicating the presence of Salmonella if carcass is improper handled. The sampling sites positive for Salmonella were also positive for PAdV. MRV was detected in 10 swabs, from only two abattoirs, mainly in outer surface of carcasses. TTSuV was also detected in all abattoirs. Our study has revealed a diverse group of viruses, each serving as indicator of either fecal (NoV, RVA, PAdV, MRV) or blood contamination (TTSuV). TTSuV could be relevant as blood contamination indicators, crucial for viruses with a viremic stage, such as HEV. The simultaneous presence of PAdV with Salmonella is relevant, suggesting PAdV as a promising indicator for fecal contamination for both bacterial and viruses. In conclusion, even in the absence of HEV, the widespread presence of Salmonella at various points in the chain, underscores the need for vigilant monitoring and mitigation strategies which could be achieved by testing not only bacteria indicators as expected by current regulation, but also some viruses (PAdV, TTSuV, MRV) which could represent other sources of fecal contamination.
Topics: Animals; Abattoirs; Feces; Food Contamination; Hepatitis E virus; Italy; Mammals; Meat; Salmonella; Swine; Viruses
PubMed: 38460205
DOI: 10.1016/j.rvsc.2024.105209 -
Frontiers in Immunology 2024Piscine red blood cells (RBC) are nucleated and have been characterized as mediators of immune responses in addition to their role in gas exchange. Salmonid RBC are...
Piscine red blood cells (RBC) are nucleated and have been characterized as mediators of immune responses in addition to their role in gas exchange. Salmonid RBC are major target cells of Piscine orthoreovirus1 (PRV-1), the etiological agent of heart and skeletal muscle inflammation (HSMI) in farmed Atlantic salmon (). PRV-1 replicates in RBC , but no viral amplification has been possible in available A. salmon cell lines. To compare RBC basal transcripts and transcriptional responses to PRV-1 in the early phase of infection with non-susceptible cells, we exposed A. salmon RBC, Atlantic salmon kidney cells (ASK) and Salmon head kidney cells (SHK-1) to PRV-1 for 24 h. The RNA-seq analysis of RBC supported their previous characterization as pluripotent cells, as they expressed a wide repertoire of genes encoding pattern recognition receptors (PRRs), cytokine receptors, and genes implicated in antiviral activities. The comparison of RBC to ASK and SHK-1 revealed immune cell features exclusively expressed in RBC, such as genes involved in chemotactic activity in response to inflammation. Differential expression analysis of RBC exposed to PRV-1 showed 46 significantly induced genes (≥ 2-fold upregulation) linked to the antiviral response pathway, including RNA-specific PRRs and interferon (IFN) response factors. In SHK-1, PRV induced a more potent or faster antiviral response (213 genes induced). ASK cells showed a differential response pattern (12 genes induced, 18 suppressed) less characterized by the dsRNA-induced antiviral pathway. Despite these differences, the RIG-I-like receptor 3 () in the family of cytosolic dsRNA receptors was significantly induced in all PRV-1 exposed cells. IFN regulatory factor 1 () was significantly induced in RBC only, in contrast to induced in SHK-1. Differences in IRF expression and activity may potentially affect viral propagation.
Topics: Animals; Salmo salar; Reoviridae Infections; Inflammation; Erythrocytes; Gene Expression Profiling; Antiviral Agents; Orthoreovirus
PubMed: 38420125
DOI: 10.3389/fimmu.2024.1359552 -
PLoS Pathogens Feb 2024Mammalian orthoreovirus (MRV) is a prototypic member of the Spinareoviridae family and has ten double-stranded RNA segments. One copy of each segment must be faithfully...
Mammalian orthoreovirus (MRV) is a prototypic member of the Spinareoviridae family and has ten double-stranded RNA segments. One copy of each segment must be faithfully packaged into the mature virion, and prior literature suggests that nucleotides (nts) at the terminal ends of each gene likely facilitate their packaging. However, little is known about the precise packaging sequences required or how the packaging process is coordinated. Using a novel approach, we have determined that 200 nts at each terminus, inclusive of untranslated regions (UTR) and parts of the open reading frame (ORF), are sufficient for packaging S gene segments (S1-S4) individually and together into replicating virus. Further, we mapped the minimal sequences required for packaging the S1 gene segment into a replicating virus to 25 5' nts and 50 3' nts. The S1 UTRs, while not sufficient, were necessary for efficient packaging, as mutations of the 5' or 3' UTRs led to a complete loss of virus recovery. Using a second novel assay, we determined that 50 5' nts and 50 3' nts of S1 are sufficient to package a non-viral gene segment into MRV. The 5' and 3' termini of the S1 gene are predicted to form a panhandle structure and specific mutations within the stem of the predicted panhandle region led to a significant decrease in viral recovery. Additionally, mutation of six nts that are conserved across the three major serotypes of MRV that are predicted to form an unpaired loop in the S1 3' UTR, led to a complete loss of viral recovery. Overall, our data provide strong experimental proof that MRV packaging signals lie at the terminal ends of the S gene segments and offer support that the sequence requirements for efficient packaging of the S1 segment include a predicted panhandle structure and specific sequences within an unpaired loop in the 3' UTR.
Topics: Animals; Orthoreovirus, Mammalian; 3' Untranslated Regions; Open Reading Frames; RNA, Viral; Mutation; Genome, Viral; Mammals
PubMed: 38394338
DOI: 10.1371/journal.ppat.1012037 -
Microbes and Infection 2024Pteropine orthoreovirus (PRV) causes respiratory tract infections in humans. Despite its emergence as a zoonotic and respiratory virus, little is known about its cell...
Pteropine orthoreovirus (PRV) causes respiratory tract infections in humans. Despite its emergence as a zoonotic and respiratory virus, little is known about its cell tropism, which hampers progress in fully understanding its pathogenesis in humans. Hek293 cells are most susceptible to PRV infection, while HeLa cells are the least. Human cytokeratin 1 (CK1) was identified as the protein that interacts with PRV. The immunofluorescence assay and qPCR results revealed prior treatment with anti-CK1 may provide Hek293 cells protection against PRV. The KRT1-knockout Hek293 cells were less susceptible to PRV infection. Further study into the pathogenesis of PRV in humans is needed.
Topics: Animals; Humans; Fish Diseases; HEK293 Cells; HeLa Cells; Keratins; Orthoreovirus; Reoviridae Infections
PubMed: 38380604
DOI: 10.1016/j.micinf.2023.105243 -
Journal of Virology Mar 2024Grass carp reovirus (GCRV), particularly the highly prevalent type II GCRV (GCRV-II), causes huge losses in the aquaculture industry. However, little is known about the...
Grass carp reovirus (GCRV), particularly the highly prevalent type II GCRV (GCRV-II), causes huge losses in the aquaculture industry. However, little is known about the mechanisms by which GCRV-II invades grass carp and further disseminates among tissues. In the present study, monocytes/macrophages (Mo/Mφs) were isolated from the peripheral blood of grass carp and infected with GCRV-II. The results of indirect immunofluorescent microscopy, transmission electron microscopy, real-time quantitative RT-PCR (qRT-PCR), western blot (WB), and flow cytometry analysis collectively demonstrated that GCRV-II invaded Mo/Mφs and replicated in them. Additionally, we observed that GCRV-II induced different types (M1 and M2) of polarization of Mo/Mφs in multiple tissues, especially in the brain, head kidney, and intestine. To assess the impact of different types of polarization on GCRV-II replication, we recombinantly expressed and purified the intact cytokines CiIFN-γ2, CiIL-4/13A, and CiIL-4/13B and successfully induced M1 and M2 type polarization of macrophages using these cytokines through experiments. qRT-PCR, WB, and flow cytometry analyses showed that M2 macrophages had higher susceptibility to GCRV-II infection than other types of Mo/Mφs. In addition, we found GCRV-II induced apoptosis of Mo/Mφs to facilitate virus replication and dissemination and also detected the presence of GCRV-II virus in plasma. Collectively, our findings indicated that GCRV-II could invade immune cells Mo/Mφs and induce apoptosis and polarization of Mo/Mφs for efficient infection and dissemination, emphasizing the crucial role of Mo/Mφs as a vector for GCRV-II infection.IMPORTANCEType II grass carp reovirus (GCRV) is a prevalent viral strain and causes huge losses in aquaculture. However, the related dissemination pathway and mechanism remain largely unclear. Here, our study focused on phagocytic immune cells, monocytes/macrophages (Mo/Mφs) in blood and tissues, and explored whether GCRV-II can invade Mo/Mφs and replicate and disseminate via Mo/Mφs with their differentiated type M1 and M2 macrophages. Our findings demonstrated that GCRV-II infected Mo/Mφs and replicated in them. Furthermore, GCRV-II infection induces an increased number of M1 and M2 macrophages in grass carp tissues and a higher viral load in M2 macrophages. Furthermore, GCRV-II induced Mo/Mφs apoptosis to release viruses, eventually infecting more cells. Our study identified Mo/Mφs as crucial components in the pathway of GCRV-II dissemination and provides a solid foundation for the development of treatment strategies for GCRV-II infection.
Topics: Animals; Apoptosis; Carps; Cytokines; Fish Diseases; Macrophages; Monocytes; Orthoreovirus; Reoviridae Infections; Virus Replication
PubMed: 38345385
DOI: 10.1128/jvi.01469-23 -
Fish & Shellfish Immunology Apr 2024The hemorrhagic disease causing by grass carp reovirus (GCRV) infection, is associated with major economic losses and significant impact on aquaculture worldwide. VP4 of...
The hemorrhagic disease causing by grass carp reovirus (GCRV) infection, is associated with major economic losses and significant impact on aquaculture worldwide. VP4 of GCRV is one of the major outer capsid proteins which can induce an immune response in the host. In this study, pNZ8148-VP4/L. lactis was constructed to express recombinant VP4 protein of GCRV, which was confirmed by the Western-Blot and enzyme-linked immunosorbent assay. Then we performed the oral immunization for rare minnow model and the challenge with GCRV-II. After oral administration, pNZ8148-VP4/L. lactis can continuously reside in the intestinal tract to achieve antigen presentation. The intestinal and spleen samples were collected at different time intervals after immunization, and the expression of immune-related genes was detected by real-time fluorescence quantitative PCR. The results showed that VP4 recombinant L. lactis could induce complete cellular and humoral immune responses in the intestinal mucosal system, and effectively regulate the immunological effect of the spleen. The immunogenicity and the protective efficacy of the oral vaccine was evaluated by determining IgM levels and viral challenge to vaccinated fish, a significant level (P < 0.01) of antigen-specific IgM with GCRV-II neutralizing activity was able to be detected, which provided a effective protection in the challenge experiment. These results indicated that an oral probiotic vaccine with VP4 expression can provide effective protection for grass carp against GCRV-II challenge, suggesting a promising vaccine strategy for fish.
Topics: Animals; Carps; Viral Vaccines; Reoviridae; Reoviridae Infections; Immunization; Orthoreovirus; Recombinant Proteins; Antibodies, Viral; Immunoglobulin M; Fish Diseases
PubMed: 38341115
DOI: 10.1016/j.fsi.2024.109439 -
Poultry Science Apr 2024Avian reovirus (ARV), which commonly induces viral arthritis or tenosynovitis and immunosuppression in chickens, is associated with the nonstructural protein p17 that...
Avian reovirus (ARV), which commonly induces viral arthritis or tenosynovitis and immunosuppression in chickens, is associated with the nonstructural protein p17 that plays a crucial role in viral replication and regulates cellular signaling pathways through its interaction with cellular proteins. In our previous study, we identified the host protein IFN-γ-inducible protein-16 (IFI16) as an interacting partner of ARV p17 through yeast two-hybrid screening. In the current study, we further confirmed the interaction between IFI16 and p17 protein using coimmunoprecipitation, glutathione S-transferase (GST)-pulldown assay, and laser confocal microscopy techniques. Additionally, we found that the amino acid of p17 is responsible for mediating the interaction with the HINa and HINb domains of IFI16. Interestingly, we observed a significant increase in IFI16 expression upon ARV infection or p17 protein exposure. Moreover, the replication of ARV was found to be largely influenced by the quantity of IFI16 protein. Overexpression of IFI16 led to a significant decrease in ARV replication, while knockdown of the IFI16 expression led to the contrary result. Additionally, our findings demonstrate that IFI16 plays a crucial role in the induction of inflammatory cytokines IFN-β and IL-1β during ARV infection as confirmed by qRT-PCR and ELISA analyses. In conclusion, our study provides novel insights into the functional role of p17 protein and the pathogenic mechanism underlying ARV infection, particularly its association with inflammatory response. Furthermore, it offers new perspectives for identifying potential therapeutic targets against ARV infection.
Topics: Animals; Chlorocebus aethiops; Orthoreovirus, Avian; Chickens; Virus Replication; Vero Cells; Immunosuppression Therapy
PubMed: 38335672
DOI: 10.1016/j.psj.2024.103506 -
Tropical Biomedicine Dec 2023Bats are flying mammals with unique immune systems that allow them to hold many pathogens. Hence, they are recognised as the reservoir of many zoonotic pathogens. In...
Bats are flying mammals with unique immune systems that allow them to hold many pathogens. Hence, they are recognised as the reservoir of many zoonotic pathogens. In this study, we performed molecular detection to detect coronaviruses, paramyxoviruses, pteropine orthoreoviruses and dengue viruses from samples collected from insectivorous bats in Krau Reserve Forest. One faecal sample from Rhinolophus spp. was detected positive for coronavirus. Based on BLASTN, phylogenetic analysis and pairwise alignment-based sequence identity calculation, the detected bat coronavirus is most likely to be a bat betacoronavirus lineage slightly different from coronavirus from China, Philippines, Thailand and Luxembourg. In summary, continuous surveillance of bat virome should be encouraged, as Krau Reserve Forest reported a wide spectrum of biodiversity of insectivorous and fruit bats. Moreover, the usage of primers for the broad detection of viruses should be reconsidered because geographical variations might possibly affect the sensitivity of primers in a molecular approach.
Topics: Animals; Coronavirus; Chiroptera; Animals, Wild; Phylogeny; Genome, Viral; Coronavirus Infections
PubMed: 38308834
DOI: 10.47665/tb.40.4.012 -
Fish & Shellfish Immunology Mar 2024Peroxiredoxins (Prxs) are a family of antioxidant enzymes crucial for shielding cells against oxidative damage from reactive oxygen species (ROS). In this study, we...
Peroxiredoxins (Prxs) are a family of antioxidant enzymes crucial for shielding cells against oxidative damage from reactive oxygen species (ROS). In this study, we cloned and analyzed two grass carp peroxiredoxin genes, CiPrx5 and CiPrx6. These genes exhibited ubiquitous expression across all sampled tissues, with their expression levels significantly modulated upon exposure to grass carp reovirus (GCRV). CiPrx5 was localized in the mitochondria, while CiPrx6 was uniformly distributed in the whole cells. Transfection or transformation of CiPrx5 and CiPrx6 into fish cells or E. coli significantly enhanced host resistance to HO and heavy metals, leading to increased cell viability and reduced cell apoptosis rates. Furthermore, purified recombinant CiPrx5 and CiPrx6 proteins effectively protected DNA against oxidative damage. Notably, overexpression of both peroxiredoxins in fish cells effectively inhibited GCRV replication, reduced intracellular ROS levels induced by GCRV infection and HO treatment, and induced autophagy. Significantly, these functions of CiPrx5 and CiPrx6 in GCRV replication and ROS mitigation were abolished upon treatment with an autophagy inhibitor. In summation, our findings suggest that grass carp Prx5 and Prx6 promote autophagy to inhibit GCRV replication, decrease intracellular ROS, and provide protection against oxidative stress.
Topics: Animals; Carps; Reactive Oxygen Species; Peroxiredoxins; Escherichia coli; Hydrogen Peroxide; Reoviridae; Reoviridae Infections; Orthoreovirus; Oxidative Stress; Autophagy; Fish Diseases
PubMed: 38301812
DOI: 10.1016/j.fsi.2024.109419