The present study describes the use of molecular methods in studying infectious salmon anaemia virus (ISAV), an important pathogen of farmed salmon in Norway, Scotland, the Faeroe Islands, Canada, USA and Chile. The nucleotide sequences of the haemagglutinin gene (HA) from 70 ISAV isolates have been analysed for phylogenetic relationship and the average mutation rate of nucleotide substitutions calculated. The isolates constitute 2 major groups, 1 European and 1 North American group. The isolate from Chile is closely related to the North American isolates. The European isolates can be further divided into 3 separate groups reflecting geographical distribution, time of collection, and transmission connected with farming activity. Based on existing information about infectious salmon anaemia (ISA) and new information emerging from the present study, it is hypothesised that: (1) ISAV is maintained in wild populations of trout and salmon in Europe; (2) it is transmitted between wild hosts mainly during their freshwater spawning phase in rivers; (3) wild salmonids, mainly trout, possibly carry benign wild-type ISAV isolates; (4) a change (mutation) in virulence probably results from deletions of amino acid segments from the highly polymorphic region (HPR) of benign wild-type isolates; (5) ISA emerges in farmed Atlantic salmon when mutated isolates are transmitted from wild salmonids or, following mutation of benign isolates, in farmed salmon after transmission from wild salmonids; (6) farming activity is an important factor in transmission of ISAV between farming sites in addition to transmission of ISAV from wild salmonids to farmed salmon; (7) transmission of ISAV from farmed to wild salmonids probably occurs less frequently than transmission from wild to farmed fish due to lower frequency of susceptible wild individuals; (8) the frequency of new outbreaks of ISA in farmed salmon probably reflects natural variation in the prevalence of ISAV in wild populations of salmonids. KEY WORDS: Infectious salmon anaemia virus · ISAV · Maintenance · Reservoir · EpizooticsResale or republication not permitted without written consent of the publisher

In the present study, 24 smolt production sites were screened for the presence of infectious salmon anaemia virus (ISAV) with the help of a specific real-time RT PCR assay, and 22 of these sites had smolts that were positive. If these smolt production sites are representative for the prevalence of ISAV in Norwegian smolts, then most marine production sites must be considered to be positive for ISAV. In addition, 92 European ISAV isolates have been genotyped based on the hemagglutinin-esterase gene (HE), and their distribution pattern was analysed. This pattern has been coupled to information about the origin of smolt, eggs, and broodfish in those cases where it has been possible to obtain such information, and with information about ISAV in neighbouring farms. The pattern suggests that an important transmission route for the ISAV could be that the salmon farming industry in Norway is circulating some of the isolates in the production cycle, i.e. some sort of vertical or transgenerational transmission may occur. It has also been shown that avirluent ISAV isolates are fairly common in Norwegian farmed salmon. Based on this, it is hypothesized that the change from avirulent to virulent ISAV isolates is a stochastic event that is dependent on the replication frequency of the virus and the time available for changes in a highly polymorphic region (HPR) of the HE gene to occur. This, and the possibility that only avirluent ISAV isolates are vertically transmitted, may explain why ISA most often occurs at marine sites and why no more than about 15 farms get ISA every year in Norway.

The phylum Chlamydiae contains obligate intracellular bacteria, several of which cause disease in their hosts. Morphological studies have suggested that this group of bacteria may be pathogens of fish, causing cysts in epithelial tissue - epitheliocystis. Recently, the first genetic evidence of a chlamydial aetiology of this disease in seawater reared Atlantic salmon from Norway and Ireland was presented, and the agent was given the name 'Candidatus Piscichlamydia salmonis'. In this article we present molecular evidence for the existence of a novel Chlamydiae that also may cause epitheliocystis in Norwegian salmonids. This novel Chlamydiae has been found in salmonid fish from freshwater, and based on its partial 16S rRNA gene, it may constitute a third genus in the family Chlamydiaceae, or a closely related sister family. By using whole-mount RNA-RNA hybridization we demonstrate how infected cells are distributed in a patchy manner on a gill arch. The morphology of the novel Chlamydiae includes the characteristic head-and-tail cells that have been described earlier from salmonid fish suffering from epitheliocystis. We propose the name 'Candidatus Clavochlamydia salmonicola' for this agent of epitheliocystis in freshwater salmonids.

Studies of infectious salmon anaemia virus (ISAV), an important pathogen of farmed salmon in Norway, Scotland, the Faeroe Islands, Ireland, Canada, the USA and Chile, suggest that natural reservoirs for this virus can be found on both sides of the North Atlantic. Based on existing information about ISAV it is believed to be maintained in wild populations of trout and salmon in Europe. It has further been suggested that ISAV is transmitted between wild hosts, mainly during their freshwater spawning phase in rivers, and that wild salmonids, mainly trout, are possible carriers of benign wild-type variants of ISAV. Change in virulence is probably a result of deletions of amino acid segments from the highly polymorphic region (HPR) of benign wild-type isolates after transmission to farmed salmon. Hence, it has been suggested that the frequency of new outbreaks of ISA in farmed salmon could partly reflect natural variation in the prevalence of ISAV in wild populations of salmonids. The aims of the present study were to screen for ISAV in wild salmonids during spawning in rivers and to determine the pathogenicity of resultant isolates from wild fish. Tissues from wild salmonids were screened by RT-PCR and real-time PCR. The prevalence of ISAV in wild trout Salmo trutta varied from 62 to 100% between tested rivers in 2001. The prevalence dropped in 2002, ranging from 13 to 36% in the same rivers and to only 6% in 2003. All ISAV were nonpathogenic when injected into disease-free Atlantic salmon, but were capable of propagation, as indicated by subsequent viral recovery. However, non-pathogenic ISAV has also been found in farmed salmon, where a prevalence as high as 60% has been registered, but with no mortalities occurring. Based on the results of the present and other studies, it must be concluded that vital information about the importance of wild and man-made reservoirs for the emergence of ISA in salmon farming is still lacking. This information can only be gained by further screening of possible reservoirs, combined with the development of a molecular tool for typing virulence and the geographical origin of the virus isolates. KEY WORDS: ISA virus · Wild salmonids · Natural reservoir Resale or republication not permitted without written consent of the publisherDis Aquat Org 66: [71][72][73][74][75][76][77][78][79] 2005 prior to the advent of intensive salmon aquaculture, suggests the long-term maintenance of ISAV in a wild reservoir. ISAV and raised levels of ISAV-specific antibodies have been found in wild salmonids with no clear relationship to aquaculture activity (Raynard et al. 2001a, K. Falk pers. comm.). Furthermore, studies based on sequencing of a so-called highly polymorphic region (HPR) of the haemagglutinin gene have suggested that all pathogenic ISAV isolates may be derived by differential deletions or by recombination of a potential ancestral sequence obtained from wild fish (Devold et al. 2001, Cunningham et al. 2002, Mjaaland et al. 2002, Nylund et al. 2003. Coupled to the fact that a r...

The microsporidian Paranucleospora theridion was discovered in Atlantic salmon Salmo salar suffering from proliferative gill disease in a marine farm in western Norway in 2008. The parasite develops in cells of the reticuloendothelial system, cells important for normal immune function. The aim of this study was to see if P. theridion could play a part in some of the diseases with unclear causes in salmon production in Norway, i.e. proliferative gill disease (PGI), pancreas disease (PD), heart and skeletal muscle inflammation (HSMI) and cardiomyopathy syndrome (CMS). P. theridion was present in all areas with salmon farming in Norway, but high prevalence and densities of the parasite in salmon and salmon lice were only seen in southern Norway. This region is also the main area for PGI and PD in Norway. Quantification of pathogens associated with PGI, PD, HSMI and CMS diagnoses showed that P. theridion levels are high in southern Norway, and may therefore play a role in susceptibility and disease development. However, among the different diagnoses, fish with PGI are particularly heavily infected with P. theridion. Therefore, P. theridion appears as a possible primary agent in cases with high mortality in connection with PGI in western Norway. KEY WORDS: Paranucleospora theridion · Atlantic salmon · Microsporidia · Proliferative gill disease · PGI · Pancreas disease · PD Resale or republication not permitted without written consent of the publisherDis Aquat Org 94: [41][42][43][44][45][46][47][48][49][50][51][52][53][54][55][56][57] 2011 Clavochlamydia salmonicola , Atlantic salmon paramyxovirus (Kvellestad et al. 2005), and salmon gill poxvirus . These agents may all contribute to the gill pathology, hypertrophia, hyperplasia, inflammation and necrosis of gill tissues, but it is not known whether they are primary pathogens or secondary invaders.It is well documented that 'pancreas disease' is associated with the presence of different strains of SAV (Nelson et al. 1995, McLoughlin et al. 1996, Villoing et al. 2000, Hodneland et al. 2005, Fringuelli et al. 2008). However, the observed disease and mortality in salmon and rainbow trout farms are different from what can be seen in challenge experiments using the different SAV strains . The explanations for this could be presence of different pathogens or other stress factors that may trigger the disease or be the primary cause of disease, reducing SAV to a secondary cause of the observed mortality in farms. Lack of comparable mortality and to a certain extent also pathology has also been observed when comparing challenge experiments using CMS and HSMI homogenates with the observed situation in salmon farms affected by these 2 diseases (Kongtorp et al. 2004a, Fritsvold et al. 2009. Hence, the trigger or primary causes for these diseases could be environmental factors or other pathogens.Paranucleospora theridion was first discovered in farmed Atlantic salmon suffering from gill disease in western Norway in (A. Nylund et al. 2009a,b, S. Nylund et al. 2009. Th...