Infectious salmon anaemia virus-molecular biology and pathogenesis of the infection.

Aquaculture has a long history in many parts of the world, but it is still young at an industrial scale. Marine fish farming in open nets of a single fish species at high densities compared to their wild compatriots opens a plethora of possible infections. Infectious salmon anaemia (ISA) is an example of disease that surfaced after large-scale farming of Atlantic salmon (Salmo salar) appeared. Here, a review of the molecular biology of the ISA virus (ISAV) with emphasis on its pathogenicity is presented. The avirulent HPR0 variant of ISAV has resisted propagation in cell cultures, which has restricted the ability to perform in vivo experiments with this variant. The transition from avirulent HPR0 to virulent HPRΔ has not been methodically studied under controlled experimental conditions, and the triggers of the transition from avirulent to virulent forms have not been mapped. Genetic segment reassortment, recombination and mutations are important mechanisms in ISAV evolution, and for the development of virulence. In the 25 years since the ISAV was identified, large amounts of sequence data have been collected for epidemiologic and transmission studies, however, the lack of good experimental models for HPR0 make the risk evaluation of the presence of this avirulent, ubiquitous variant uncertain. This review summarizes the current knowledge related to molecular biology and pathogenicity of this important aquatic orthomyxovirus.

Identification and isolation of infective filamentous particles in Infectious Salmon Anemia Virus (ISAV).

The infectious salmon anemia virus (ISAV) is an aquatic pathogen that is a member of the Orthomyxoviridae family with lethal hemorrhagic potential. Although it affects other species of salmonid fish, ISAV only causes disease in Atlantic salmon (Salmo salar) specimens in sea water. In spite of the fact that the virus has been described as enveloped with icosahedral symmetry, viral like particles with anomalous morphology have been observed in field samples, this we have not been able to recover then in adequate quantities for full demonstration. We report a procedure to concentrate and recover these novel forms of the virus, comparing two cell lines from different origins, demonstrating that these forms were preferentially expressed in cells of epithelial origin.

Genetic analysis and comparative virulence of infectious salmon anemia virus (ISAV) types HPR7a and HPR7b from recent field outbreaks in Chile.

BACKGROUND: Infectious salmon anemia (ISA) is a serious disease of marine farmed Atlantic salmon, Salmo salar L. caused by ISA virus (ISAV). ISAV genomic segments 5 and 6 encode surface glycoproteins hemagglutinin-esterase (HE) and F protein important for the pathogenicity of ISAV. In this study, we describe the genetic characteristics and relationship between ISAV-HPR7a and ISAV-HPR7b strains that caused the ISA outbreaks in Chile in 2013 and 2014, respectively, and the evolution of the ISAV clades since 2009 based on segment 5 and 6 sequences. METHODS: The study material included samples from six ISA cases in Chile. RNA was extracted from salmon tissues and ISAV isolated from cell culture; segments 5 and 6 were amplified by RT-PCR and compared by alignment with ISAV sequences from the GenBank database. RESULTS: ISAV-HPR7a and ISAV-HPR7b belong to the European Genotype I strains only found in Europe and Chile, and in both cases, show high similarity in segments 5 and 6 with identity between 95-96%. Our data confirm the hypothesis that the original virus was introduced to Chile in 1996. Compared to the 2007 ISAV-HPR7b isolate, the 2014 ISAV-HPR7b does not have an insertion in segment 5 and was associated with low mortality, which suggests that ISAV virulence was attenuated by the absence of the insertion in segment 5. In contrast, the highly virulent ISAV-HPR14 from April 2013 outbreak did not have the insertion in segment 5 either. CONCLUSION: Variability in the ISAV virulence markers supports the quasispecies theory that multiple evolution forces are likely to shape ISAV genetic diversity. Our findings provide evidence of continuing evolution of ISAV in the Chilean aquaculture industry.

Infectious salmon anaemia - pathogenesis and tropism.

Infectious salmon anaemia (ISA) is a serious disease of farmed Atlantic salmon caused by the aquatic orthomyxovirus infectious salmon anaemia virus (ISAV). ISA was first detected in Norway in 1984 and was characterized by severe anaemia and circulatory disturbances. This review elucidates factors related to the pathogenesis of ISA in Atlantic salmon, the dissemination of the virus in the host and the general distribution of the 4-O-acetylated sialic acids ISAV receptor. The knowledge contributes to the understanding of this disease, and why, almost 30 years after the first detection, it is still causing problems for the aquaculture industry.

Infectious salmon anaemia virus infection of Atlantic salmon gill epithelial cells.

Infectious salmon anaemia virus (ISAV), a member of the Orthomyxoviridae family, infects and causes disease in farmed Atlantic salmon (Salmo salar L.). Previous studies have shown Atlantic salmon endothelial cells to be the primary targets of ISAV infection. However, it is not known if cells other than endothelial cells play a role in ISAV tropism. To further assess cell tropism, we examined ISAV infection of Atlantic salmon gill epithelial cells in vivo and in vitro. We demonstrated the susceptibility of epithelial cells to ISAV infection. On comparison of primary gill epithelial cell cultures with ISAV permissive fish cell cultures, we found the virus yield in primary gill epithelial cells to be comparable with that of salmon head kidney (SHK)-1 cells, but lower than TO or Atlantic salmon kidney (ASK)-II cells. Light and transmission electron microscopy (TEM) revealed that the primary gill cells possessed characteristics consistent with epithelial cells. Virus histochemistry showed that gill epithelial cells expressed 4-O-acetylated sialic acid which is recognized as the ISAV receptor. To the best of our knowledge, this is the first demonstration of ISAV infection in Atlantic salmon primary gill epithelial cells. This study thus broadens our understanding of cell tropism and transmission of ISAV in Atlantic salmon.

Modelling the spread of infectious salmon anaemia among salmon farms based on seaway distances between farms and genetic relationships between infectious salmon anaemia virus isolates.

Infectious salmon anaemia (ISA) is an important infectious disease in Atlantic salmon farming causing recurrent epidemic outbreaks worldwide. The focus of this paper is on tracing the spread of ISA among Norwegian salmon farms. To trace transmission pathways for the ISA virus (ISAV), we use phylogenetic relationships between virus isolates in combination with space-time data on disease occurrences. The rate of ISA infection of salmon farms is modelled stochastically, where seaway distances between farms and genetic distances between ISAV isolates from infected farms play prominent roles. The model was fitted to data covering all cohorts of farmed salmon and the history of all farms with ISA between 2003 and summer 2009. Both seaway and genetic distances were significantly associated with the rate of ISA infection. The fitted model predicts that the risk of infection from a neighbourhood infectious farm decreases with increasing seaway distance between the two farms. Furthermore, for a given infected farm with a given ISAV genotype, the source of infection is significantly more likely to be ISAV of a small genetic distance than of moderate or large genetic distances. Nearly half of the farms with ISA in the investigated period are predicted to have been infected by an infectious farm in their neighbourhood, whereas the remaining half of the infected farms had unknown sources. For many of the neighbourhood infected farms, it was possible to point out one or a few infectious farms as the most probable sources of infection. This makes it possible to map probable infection pathways.

Transcriptomic analysis of responses to infectious salmon anemia virus infection in macrophage-like cells.

The aquatic orthomyxovirus infectious salmon anemia virus (ISAV) is an important pathogen for salmonid aquaculture, however little is known about protective and pathological host responses to infection. We have investigated intracellular responses during cytopathic ISAV infection in the macrophage-like Atlantic salmon kidney (ASK) cell line by microarray analysis (1.8k SFA2.0 immunochip) and a functional assay for glutathione. Gene transcription changed rapidly and consistently with time and with minor differences between two virus isolates. While several pro-inflammatory and antiviral immune genes were induced, genes involved in cell signaling and integrity were down-regulated, suggesting isolation of infected cells from cell-to-cell interaction and responses to external signals. Differential expression of genes regulating cell cycle and apoptosis implied opposite cues from host cell and virus. This was in pace with massive down-regulation of genes involved in biosynthesis and processing of nucleotides and nucleic acids. Significant down-regulation of several genes involved in metabolism of reactive oxygen species suggested increased oxidative stress, which was confirmed by a functional assay showing reduced levels of glutathione during infection. Testing of expression data against a microarray database containing diverse experiments revealed candidate marker genes for ISAV infection. Our findings provide novel insight into cellular host responses and determinants for acute cytopathic ISAV infection.

Morphologic description of infectious salmon anaemia virus (ISAV)-induced lesions in rainbow trout Oncorhynchus mykiss compared to Atlantic salmon Salmo salar.

In the present study the pathogenesis of experimental infectious salmon anaemia virus (ISAV) infection in rainbow trout Oncorhynchus mykiss (Walbaum, 1972) and Atlantic salmon Salmo salar was compared. The virus infection in the 2 species demonstrated different mortality patterns and pathology characteristics. Atlantic salmon showed a typical acute mortality pattern peaking at 8 to 16 d post-infection (dpi) depending on virus dose, whereas in rainbow trout, only the highest virus dose (10(7.13-7.8) TCID50/200 microl) showed a similar pattern. The middle (10(4.13) TCID50/200 microl) and lowest virus doses (10(2.13) TCID50/200 microl) in rainbow trout induced only sporadic protracted mortality, lasting up to 46 dpi. Infected rainbow trout that were live-sampled and those that died demonstrated increased erythrophagia, clusters of cellular degeneration in the haematopoietic portion of the kidney, and occasionally epicarditis, endocarditis and myocarditis. These lesions are very different from the typical necrosis in liver and kidney that occur in infected Atlantic salmon, and some of them may be indicative of an antiviral response by a resistant host to the ISAV infection. Virus was detected in the endothelium of the rainbow trout tissues using in situ hybridization, supporting our conclusions of the ISAV-induced lesions in this report.

Hydrographics and the timing of infectious salmon anemia outbreaks among Atlantic salmon (Salmo salar L.) farms in the Quoddy region of Maine, USA and New Brunswick, Canada.

Infectious salmon anemia (ISA) has caused severe morbidity and mortality in farmed Atlantic salmon in North America, Norway, Scotland and the Faroe Islands. The Quoddy region of Maine, United States of America (USA), and New Brunswick (NB), Canada is characterized by extensive tidal mixing and close proximity between farms. This region is also prone to recurrent appearances of ISA, though control measures limit disease spread and severity on infected farms. We conducted a retrospective longitudinal analysis of the apparent impact of hydrographics on the incidence and timing of ISA outbreaks on Atlantic salmon (Salmo salar L.) farms in the Quoddy region from May 2002 to August 2004. A time-series cross-sectional regression of 32 farms over 28 months demonstrated a limited, but statistically significant, spatio-temporal clustering of ISA outbreaks linked hydrographically. New outbreaks correlated temporally with those occurring on-site 1 and 3 months prior, and those occurring within one tidal-excursion upstream the same month. Other risk factors included holdover of previous year-class fish, wharf sharing, and possibly harvests of cages infected in previous months. Conclusions suggest that tidal dispersion does play a role in ISAV transmission in the Quoddy region. Dispersal of free virus and/or tidal distribution of lice or other hydrographically influenced vectors or fomites could all contribute to the spatio-temporal patterns described.