Dual Mutation Events in the Haemagglutinin-Esterase and Fusion Protein from an Infectious Salmon Anaemia Virus HPR0 Genotype Promote Viral Fusion and Activation by an Ubiquitous Host Protease

Funding: The Scottish Government funded this work, as part of their global budget on aquaculture research. The funder had no role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript.Peer reviewedPublisher PD

Detection of the myxosporean parasite Parvicapsula pseudobranchicola in Atlantic salmon (Salmo salar L.) using in situ hybridization (ISH)

-Parvicapsula pseudobranchicola is a marine myxosporean parasite infecting farmed Atlantic salmon (Salmo salar). A major site for the parasite is the pseudobranch, which may be destroyed in heavily infected fish. Parvicapsulosis may be associated with significant mortality, although the main effect of infections seems to be runting. In situ hybridization (ISH) is, in the absence of specific antibodies, the preferred method for the detection of cell- and tissue tropisms of myxozoans in the early phases of infection of the host, and provides information about the possible association between the pathogen and pathology. A positive diagnosis of parvicapsulosis is based on histopathology and PCR. The aim of the present work was to develop a specific, sensitive and robust ISH assay for the detection of P. pseudobranchicola in tissues

Evolutionary mechanisms involved in the virulence of infectious salmon anaemia virus (ISAV), a piscine orthomyxovirus

AbstractInfectious salmon anaemia virus (ISAV) is an orthomyxovirus causing a multisystemic, emerging disease in Atlantic salmon. Here we present, for the first time, detailed sequence analyses of the full-genome sequence of a presumed avirulent isolate displaying a full-length hemagglutinin-esterase (HE) gene (HPR0), and compare this with full-genome sequences of 11 Norwegian ISAV isolates from clinically diseased fish. These analyses revealed the presence of a virulence marker right upstream of the putative cleavage site R267 in the fusion (F) protein, suggesting a Q266→L266 substitution to be a prerequisite for virulence. To gain virulence in isolates lacking this substitution, a sequence insertion near the cleavage site seems to be required. This strongly suggests the involvement of a protease recognition pattern at the cleavage site of the fusion protein as a determinant of virulence, as seen in highly pathogenic influenza A virus H5 or H7 and the paramyxovirus Newcastle disease virus

Molecular and antigenic characterization of Piscine orthoreovirus (PRV) from rainbow trout (Oncorhynchus mykiss)

Piscine orthoreovirus (PRV-1) causes heart and skeletal muscle inflammation (HSMI) in farmed Atlantic salmon (Salmo salar). Recently, a novel PRV (formerly PRV-Om, here called PRV-3), was found in rainbow trout (Oncorhynchus mykiss) with HSMI-like disease. PRV is considered to be an emerging pathogen in farmed salmonids. In this study, molecular and antigenic characterization of PRV-3 was performed. Erythrocytes are the main target cells for PRV, and blood samples that were collected from experimentally challenged fish were used as source of virus. Virus particles were purified by gradient ultracentrifugation and the complete coding sequences of PRV-3 were obtained by Illumina sequencing. When compared to PRV-1, the nucleotide identity of the coding regions was 80.1%, and the amino acid identities of the predicted PRV-3 proteins varied from 96.7% (λ1) to 79.1% (σ3). Phylogenetic analysis showed that PRV-3 belongs to a separate cluster. The region encoding σ3 were sequenced from PRV-3 isolates collected from rainbow trout in Europe. These sequences clustered together, but were distant from PRV-3 that was isolated from rainbow trout in Norway. Bioinformatic analyses of PRV-3 proteins revealed that predicted secondary structures and functional domains were conserved between PRV-3 and PRV-1. Rabbit antisera raised against purified virus or various recombinant virus proteins from PRV-1 all cross-reacted with PRV-3. Our findings indicate that despite different species preferences of the PRV subtypes, several genetic, antigenic, and structural properties are conserved between PRV-1 and-3

Infeksjoner med parasitten Nucleospora cyclopteri (Microsporidia) i rognkjeks, Cyclopterus lumpus

Source at https://www.vetinst.no/rapporter-og-publikasjoner/rapporter/2019/infeksjoner-med-parasitten-nucleospora-cyclopteri-microsporidia-i-rognkjeks-cyclopterus-lumpus.Nucleospora cyclopteri (Microsporidia) is one of many parasites infecting lumpfish, Cyclopterus lumpus, and has been shown to cause disease and mortality in lumpfish. Infections in fish are often multifactorial and the impact of one agent on the development of disease can be difficult to elucidate. In addition to mortality, infections in lumpfish can lead to diseases with subsequently lowered appetite. This is of particular importance since lumpfish are used as a biological control agent, eating salmon lice, Lepeophtherius salmonis, off the salmon. Knowledge on the different disease agents of lumpfish is therefore of utmost importance. The main aim of this project was to identify how to obtain an infection-free lumpfish in land-based hatcheries and to study the impact that N. cyclopteri has on the health of the lumpfish and thereby its effect as a biological control agent. The project therefore aimed to map the presence of N. cyclopteri and other disease agents in wild caught lumpfish and in eggs/sperm, in fry and in farmed lumpfish stocked in the sea. In addition, we wanted to study the transmission pathways and clinical significance of the parasite. Unfortunately, we were not able to obtain a group of lumpfish fry infected with N. cyclopteri that we intended to follow through the land phase. The study of pathogenesis, infection dynamics, or whether an infection with N. cyclopteri pre-disposes for secondary infections, was therefore abandoned. We studied the presence of co-infections, methods for optimal sampling and tissue tropism in wild caught lumpfish in this project. Nucleospora cyclopteri was present in 60% of the sampled individuals from the waters around Averøy, in county Møre og Romsdal. The fish were analysed with regard to a range of infectious agents (viruses, bacteria and parasites) commonly found in other fish species, or previously recorded in lumpfish. No viral agents or other important pathogens were detected, but supposedly nonpathogenic microparasites, like Kudoa islandica (Myxozoa) in the muscle tissue and coccidians in the intestine, were frequently found. Nucleospora cyclopteri was detected in all tissues examined: anterior, mid and posterior kidney, spleen, heart, gills, brain, muscle liver and blood, thus indicating that the infection is systemic. The density of N. cyclopteri was highest in the anterior kidney, followed by mid and posterior kidney, spleen and gills, while the prevalence was highest in the ventricle of the heart. Observations from this study indicate that the parasite is released through urine and bile. We also show that N. cyclopteri can be detected using swabs from the skin, gill and vent, and by blood samples and gill biopsies, thus demonstrating the possibility of non-lethal detection of N. cyclopteri in lumpfish. Amongst these, the most promising non-lethal samples for detection were gill biopsies and leukocyte fractions from blood samples. Images normal histology and pathological agents from this project is included in an openly available online image database. This image database can be accessed by diagnosticians and researchers and used when evaluating pathological findings in lumpfish. While vertical transmission cannot be excluded, the results from this project indicate that this is not the dominant route. It is in any case advisable to routinely screen broodfish for N. cyclopteri to avoid using positive individuals for the production of eggs and fry. Given that N. cyclopteri undoubtedly destroys leukocytes in high numbers and over large areas of tissue, it is reasonable to assume that the parasite has an effect on the immune competence of the fish

Catching the fish with the worm: A case study on eDNA detection of the monogenean parasite Gyrodactylus salaris and two of its hosts, Atlantic salmon (Salmo salar) and rainbow trout (Oncorhynchus mykiss)

Background: Environmental DNA (eDNA) monitoring is growing increasingly popular in aquatic systems as a valuable complementary method to conventional monitoring. However, such tools have not yet been extensively applied for metazoan fish parasite monitoring. The fish ectoparasite Gyrodactylus salaris, introduced into Norway in 1975, has caused severe damage to Atlantic salmon populations and fisheries. Successful eradication of the parasite has been carried out in several river systems in Norway, and Atlantic salmon remain infected in only seven rivers, including three in the Drammen region. In this particular infection region, a prerequisite for treatment is to establish whether G. salaris is also present on rainbow trout upstream of the salmon migration barrier. Here, we developed and tested eDNA approaches to complement conventional surveillance methods. Methods: Water samples (2 × 5 l) were filtered on-site through glass fibre filters from nine locations in the Drammen watercourse, and DNA was extracted with a CTAB protocol. We developed a qPCR assay for G. salaris targeting the nuclear ribosomal ITS1 region, and we implemented published assays targeting the mitochondrial cytochrome-b and NADH-regions for Atlantic salmon and rainbow trout, respectively. All assays were transferred successfully to droplet digital PCR (ddPCR). Results: All qPCR/ddPCR assays performed well both on tissue samples and on field samples, demonstrating the applicability of eDNA detection for G. salaris, rainbow trout and Atlantic salmon in natural water systems. With ddPCR we eliminated a low cross-amplification of Gyrodactylus derjavinoides observed using qPCR, thus increasing specificity and sensitivity substantially. Duplex ddPCR for G. salaris and Atlantic salmon was successfully implemented and can be used as a method in future surveillance programs. The presence of G. salaris eDNA in the infected River Lierelva was documented, while not elsewhere. Rainbow trout eDNA was only detected at localities where the positives could be attributed to eDNA release from upstream land-based rainbow trout farms. Electrofishing supported the absence of rainbow trout in all of the localities