PRV-1 Infected Macrophages in Melanized Focal Changes in White Muscle of Atlantic Salmon (Salmo salar) Correlates With a Pro-Inflammatory Environment

Melanized focal changes in white skeletal muscle of farmed Atlantic salmon, “black spots”, is a quality problem affecting on average 20% of slaughtered fish. The spots appear initially as “red spots” characterized by hemorrhages and acute inflammation and progress into black spots characterized by chronic inflammation and abundant pigmented cells. Piscine orthoreovirus 1 (PRV-1) was previously found to be associated with macrophages and melano-macrophages in red and black spots. Here we have addressed the inflammatory microenvironment of red and black spots by studying the polarization status of the macrophages and cell mediated immune responses in spots, in both PRV-1 infected and non-infected fish. Samples that had been collected at regular intervals through the seawater production phase in a commercial farm were analyzed by multiplex fluorescent in situ hybridization (FISH) and RT-qPCR methods. Detection of abundant inducible nitric oxide synthase (iNOS2) expressing M1-polarized macrophages in red spots demonstrated a pro-inflammatory microenvironment. There was an almost perfect co-localization with the iNOS2 expression and PRV-1 infection. Black spots, on the other side, had few iNOS2 expressing cells, but a relatively high number of arginase-2 expressing anti-inflammatory M2-polarized macrophages containing melanin. The numerous M2-polarized melano-macrophages in black spots indicate an ongoing healing phase. Co-localization of PRV-1 and cells expressing CD8+ and MHC-I suggests a targeted immune response taking place in the spots. Altogether, this study indicates that PRV-1 induces a pro-inflammatory environment that is important for the pathogenesis of the spots. We do not have indication that infection of PRV-1 is the initial causative agent of this condition.

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Pilchard orthomyxovirus (POMV). II. Causative agent of salmon orthomyxoviral necrosis, a new disease of farmed Atlantic salmon Salmo salar

Diseases of Aquatic Organisms ◽

10.3354/dao03469 ◽

2020 ◽

Vol 139 ◽

pp. 51-68 ◽

Cited By ~ 3

Author(s):

SE Godwin ◽

RN Morrison ◽

G Knowles ◽

MC Cornish ◽

D Hayes ◽

...

Keyword(s):

Cell Culture ◽

Atlantic Salmon ◽

Infected Fish ◽

Direct Role ◽

Multiple Organs ◽

Direct Exposure ◽

Salmon Fry ◽

Infected Cells ◽

Farmed Atlantic Salmon

Since 2012, an orthomyxo-like virus has been consistently linked to epizootics in marine farmed Atlantic salmon in Tasmania, Australia. Here we describe the properties of the virus, designated the pilchard orthomyxovirus (POMV), in cell culture and present data verifying its direct role in a disease of Atlantic salmon. In infected cells, viral RNA was detectable in both the nucleus and cytoplasm, consistent with the replication cycle of an orthomyxovirus. Viral replication in vitro was temperature-dependent (within a range of 10-20°C), and yields of virus were typically in excess of 107 TCID50 ml-1. In controlled infection trials, cell culture-derived POMV produced significant morbidity in Atlantic salmon fry, pre-smolt and post-smolt. In all cases, the development of disease was rapid, with moribund fish detected within 5 d of direct exposure to POMV, and maximum cumulative morbidity occurring within 4 wk. The experimentally infected fish developed a characteristic suite of gross and microscopic pathological changes, which were consistent with those observed in Atlantic salmon overtly affected by POMV-associated disease on sea farms. These included necrotic lesions across multiple organs that were directly associated with the presence of the virus. Together, our observations indicate that POMV is an endemic virus likely transmitted from wild fish to farmed Atlantic salmon in Tasmania. The virus is pathogenic to Atlantic salmon in freshwater and marine environments and causes a disease that we have named salmon orthomyxoviral necrosis.

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Dissemination of Piscine orthoreovirus-1 (PRV-1) in Atlantic Salmon (Salmo salar) during the Early and Regenerating Phases of Infection

Pathogens ◽

10.3390/pathogens9020143 ◽

2020 ◽

Vol 9 (2) ◽

pp. 143 ◽

Cited By ~ 3

Author(s):

Kannimuthu Dhamotharan ◽

Håvard Bjørgen ◽

Muhammad Salman Malik ◽

Ingvild B. Nyman ◽

Turhan Markussen ◽

...

Keyword(s):

Atlantic Salmon ◽

Salmo Salar ◽

Cellular Localization ◽

Heart Tissue ◽

Protein Levels ◽

Atlantic Salmon Salmo Salar ◽

Histopathologic Evaluation ◽

Farmed Atlantic Salmon ◽

Heart Lesions

Piscine orthoreovirus-1 (PRV-1) can cause heart and skeletal muscle inflammation (HSMI) in farmed Atlantic salmon (Salmo salar), but the line of events from infection, pathologic change, and regeneration has not been thoroughly described. In this study, the cellular localization and variation of PRV-1 RNA and protein levels were analyzed at different times post-exposure in experimentally infected Atlantic salmon. Immunohistochemistry, flow cytometry, and Western blot were used for assessment of the presence of the PRV-1 σ1 protein, while RT-qPCR and in situ hybridization were performed for viral RNA. Histopathologic evaluation demonstrated that PRV-1 infection induced heart lesions typical of HSMI, such as severe epicarditis and myocarditis with degeneration of cardiomyocytes, necrosis, and diffuse cellular infiltration. PRV-1 infection of erythrocytes and the peak viral plasma level preceded virus presence in cardiomyocytes and hepatocytes. Arginase-2-positive, macrophage-like cells observed in the heart indicated possible polarization to M2 macrophages and the onset of regenerative processes, which may contribute to the recovery from HSMI. The virus was cleared from regenerating heart tissue and from hepatocytes, but persisted in erythrocytes.

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Assessment of acute effects of in situ net cleaning on gill health of farmed Atlantic salmon (Salmo salar L)

Aquaculture ◽

10.1016/j.aquaculture.2021.737203 ◽

2021 ◽

pp. 737203

Author(s):

Liv Østevik ◽

Marit Stormoen ◽

Ane Nødtvedt ◽

Marta Alarcón ◽

Kai-Inge Lie ◽

...

Keyword(s):

Atlantic Salmon ◽

Salmo Salar ◽

Acute Effects ◽

Atlantic Salmon Salmo Salar ◽

Farmed Atlantic Salmon

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Faculty Opinions recommendation of Effects of nitric oxide synthase inhibition by L-NAME on oxygen uptake kinetics in isolated canine muscle in situ.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1087541.540599 ◽

2007 ◽

Author(s):

Michael Reid

Keyword(s):

Nitric Oxide ◽

Nitric Oxide Synthase ◽

Oxygen Uptake ◽

Oxygen Uptake Kinetics ◽

Uptake Kinetics ◽

Oxide Synthase

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Effect of vaccines against pancreas disease in farmed Atlantic salmon

Journal of Fish Diseases ◽

10.1111/jfd.13505 ◽

2021 ◽

Author(s):

Magnus Vikan Røsæg ◽

Ragnar Thorarinsson ◽

Arnfinn Aunsmo

Keyword(s):

Atlantic Salmon ◽

Pancreas Disease ◽

Farmed Atlantic Salmon

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Searching for the sweet spot of amoebic gill disease of farmed Atlantic salmon: the potential role of glycan-lectin interactions in the adhesion of Neoparamoeba perurans

International Journal for Parasitology ◽

10.1016/j.ijpara.2020.11.009 ◽

2021 ◽

Author(s):

P.C. Lima ◽

L. Hartley-Tassell ◽

O. Cooper ◽

J.W. Wynne

Keyword(s):

Atlantic Salmon ◽

Potential Role ◽

Sweet Spot ◽

Gill Disease ◽

Farmed Atlantic Salmon

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823 CD4 T cells are essential for an anti-tumor effect in a B78 murine melanoma tumor model

Journal for ImmunoTherapy of Cancer ◽

10.1136/jitc-2020-sitc2020.0823 ◽

2020 ◽

Vol 8 (Suppl 3) ◽

pp. A873-A873

Author(s):

Arika Feils ◽

Mackenzie Heck ◽

Anna Hoefges ◽

Peter Carlson ◽

Luke Zangl ◽

...

Keyword(s):

T Cells ◽

Tumor Growth ◽

Cd4 T Cells ◽

Tumor Response ◽

Immune Cell ◽

Cd8 T Cell ◽

Mhc I ◽

Mhc Ii ◽

Flow Cytometric

BackgroundMice bearing B78 melanoma tumors can be cured using an in situ vaccine (ISV) regimen that includes radiation (RT) together with immunocytokine (tumor-targeting mAb conjugated to IL-2). B78 melanoma cells, derived from B16 cells, express minimal to no MHC-I but express MHC-II upon IFN-g/TNF-a stimulation. Although B78 cells are primarily MHC-I-deficient, an increased CD8 T cell infiltration into the tumor microenvironment (TME) has been shown following ISV.1 To further investigate the potential role of specific immune cell lineages in the B78 anti-tumor response to ISV, immune subset depletion studies and flow cytometric analyses were performed.MethodsC57BL/6 mice bearing B78 tumors were depleted of immune cell subsets with mAbs (anti-CD4, anti-CD8, anti-NK1.1, or Rat IgG control) for 3 weeks during the course of treatment. Treatment groups included no treatment, RT (12 Gy), or ISV (RT D0 and immunocytokine D5-D9). 6 mice/group (repeated three times) were followed for survival/tumor growth, and flow cytometry studies included 4 mice/group, sacrificed on D8 and D13 following the start of ISV.ResultsMice depleted of CD4 T cells during the course of ISV showed a significant reduction of anti-tumor effect as compared to mice treated with ISV/Rat IgG (pConclusionsThese studies suggest that CD4 T cells are essential for an anti-tumor response in the B78 melanoma model. In vivo depletion data show that CD4 T cells, but not CD8 or NK cells, are required for a decrease in tumor growth via ISV. Flow cytometric analyses suggest an interplay between CD4 and CD8 T cells as indicated by a decrease in CD8/IFN-g expression following ISV in the absence of CD4 T cells. The role that MHC-I and MHC-II expression plays in this CD4/CD8 T cell anti-tumor response is under investigation. In future studies, B78 melanoma may serve as a critical syngeneic model for development of more effective immunotherapy treatment regimens.Ethics ApprovalAll animal experiments were performed in accordance with protocols approved by Animal Care and Use Committees of the University of Wisconsin-Madison.ReferenceMorris Z, Guy E, Francis D, et al. In situ tumor vaccination by combining local radiation and tumor-specific antibody or immunocytokine treatments. Cancer Res 2016;76(13):3929-3941.

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