The commensal microbiota plays an important role in the well-being of the host organism, and it would be worthwhile to know the tenacious communities among them. Therefore, a study was undertaken to examine the changes in constitution of the intestinal microbiota of wild fish consequential to captivity. At first, the composition of intestinal microorganisms of Atlantic cod caught from the coastal area off Bodø, Norway, was examined. Thereafter, the changes in the bacterial community of the captive fish after offering them artificial feed or subjecting them to starvation were studied. The microbiota from the intestinal contents and wall segments were analyzed quantitatively by spread plate technique and DAPI staining and qualitatively by denaturing gradient gel electrophoresis. The study revealed that the counts of intestinal microbes in wild-caught Atlantic cod were not affected by captive rearing for 6 weeks, either when fed or when starved. However, the diversity of intestinal bacterial community was reduced in response to artificial feeding, whereas the change was restricted upon starvation.

An outbreak of nodavirus infection in turbot larvae is described with respect to histopathology, immunohistochemistry, cell culture cultivation, RT-PCR amplification and sequence analysis of the capsid protein gene RNA2. Affected turbot developed classical signs of viral encephalopathy and retinopathy (VER) with abnormal swimming behaviour and high mortality levels. In the acute stage of infection, light microscopy revealed vacuolation of the central nervous system (CNS), with positive immunohistochemical staining for nodavirus. Later in the infection, CNS lesions appeared more chronic and contained clusters of cells immunopositive for nodavirus. Bacterial overgrowth in the intestines of the fish may have provoked or influenced the course of the nodavirus infection. We were unable to propagate the virus in cell culture. While RT-PCR using primers designed to detect Atlantic halibut nodavirus gave negative results, further testing with primers complementary to a more conserved region of RNA2 resulted in amplification of a product of the expected size. The entire RNA2 segment was cloned and sequenced. Sequence alignment showed that the turbot nodavirus (TNV) was different from previously described fish nodaviruses. In addition, phylogenetic analysis based on an 823 nt region of the sequence indicated that TNV clustered outside the four established fish nodavirus genotypes, suggesting a fifth genotype within the betanodaviruses.

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Homogenate of tissue from juveniles of Atlantic halibut Hippoglossus hippoglossus suffering from viral encephalopathy and retinopathy (VER) was used to challenge smolt of Atlantic salmon Salmo salar with an initial average weight of 110 g. The nodavirus was administered in the form of an intraperitoneal injection, and the fish were kept for 134 d post challenge. Genotype characterisation of the nodavirus was performed by sequencing the RNA1 and RNA2 segments, and a quantitative real-time PCR (Q-PCR) assay was developed. Tissues from different organs were stained by immunohistochemistry (IHC). Samples were collected at random on Days 7, 25, 45, 69, 125 and 134 after challenge. Mortality, clinical signs and pathology of VER were observed only in the challenged group. The Q-PCR detected positive fish only in the challenged group, all of which were positive on all days of sampling. An increase in relative virus concentrations was observed from Day 7 to Day 25 post challenge. The increased level of virus concentration was maintained in the medulla oblongata throughout the experiment, suggesting persistence or slow elimination of the virus over time. The IHC detected positive cells on Days 34, 70 and 74. These results suggest that the nodavirus is transported to the medulla oblongata from the intraperitoneal injection site and is able to replicate in salmon. When injected, this nodavirus isolate caused mortality and established a persistent infection in the challenged salmon throughout the experiment. This susceptibility suggests that co-location of salmon and marine species should be avoided until further studies of possible transmission have been carried out. KEY WORDS: Nodavirus · Atlantic salmon · Atlantic halibut · Experimental infection · Q-PCR · Immunohistochemistry · Viral encephalopathy · RetinopathyResale or republication not permitted without written consent of the publisher Dis Aquat Org 68: 7-15, 2005 such as cod Gadus morhua (Starkey et al. 2001, Johnson et al. 2002, Gagne et al. 2004, Atlantic halibut Hippoglossus hippoglossus (Grotmol et al. 1995, Grotmol et al. 1997b) and spotted wolffish Anarhichas minor , Sommer et al. 2004. In Norway, Atlantic halibut hatcheries have suffered several outbreaks of VER during the past few years (Dannevig et al. 2000, Johansen et al. 2004a, suggesting the persistence of virus in cultured populations. Although VER has been considered to be a disease of juveniles, individuals weighing several kg have also shown clinical signs of VER (Aspehaug et al. 1999). This observed persistence of nodavirus in adult fish suggests that the virus could be present in farmed halibut for a long time and may represent a risk of spreading nodavirus with movement of persistently infected animals. Transportation of infected animals into areas with potential susceptible hosts may represent one important way of spreading diseases. In Norway, Atlantic salmon Salmo salar is the dominant farmed species, but farming of marine fish such as Atlantic cod and halibut is growing. Although...

Betanodaviruses have been isolated and detected in both farmed and wild fish species worldwide. They are classified in five clusters, and all are connected to mortalities in farmed fish. The clusters do not represent specific geographical areas or host species, but one cluster, barfin flounder nervous necrosis virus (BFNNV), is mainly associated with cold water fish species. This study presents the first species-specific clade within the BFNNV cluster. This clade consists of six isolates from wild and farmed Atlantic cod in Norway and is genetically distinct from other betanodaviruses in the North Atlantic. Screening of farmed and wild cod in Norway shows that betanodaviruses are present in wild fish on the west coast of Norway, including migratory cod, but so far we have not detected any betanodavirus-positive wild cod in northern Norway. The presence of significant amounts of betanodaviruses in wild cod represents a serious challenge for the management of viral nervous necrosis in farmed cod in Norway. Betanodavirus-positive farmed cod were present both in western and northern Norway. Mortalities in three cod farms were suspected to be caused by betanodaviruses; however, in two of these, other pathogens may have been responsible for or strongly contributed to the mortalities.

Atlantic cod, Gadus morhua L., larvae were fed rotifers, Brachionus plicatilis and Artemia franciscana enriched on four different commercial media, using the manufacturers' protocols. Pooled samples of 20 cod larvae were homogenized, diluted, and plated out on Petri dishes. The number of colony-forming units per larva was estimated, and the dominant strains subsequently sampled for sequencing of 16S rDNA. Bacteria showing high sequence similarity to a pathogen characteristic of cod and other fish species, Listonella anguillarum, were present in all four groups. Other taxa present among the dominating bacterial colonies were Pseudoalteromonas sp., and Vibrio sp. However, these bacteria could be assigned to genera only. The different enrichments probably affected the number of colony-forming bacteria per millilitre in the enrichment cultures as well as in the larval gastrointestinal (GI) tract. Also, the composition of the microbiota associated with the larval GI tract was probably affected by the enrichment media.

Viral encephalopathy and retinopathy (VER) was diagnosed in 5 to 24 g sized farmed Atlantic cod Gadus morhua kept in sea cages at Parisvatn, Hordaland county, on the west coast of Norway. Moderate mortality (10 to 15%) was observed, along with anorexia and abnormal swimming behaviour, such as looping or spiral swimming and reduced coordination. Nodavirus was detected by 2 different real-time RT-PCR assays, and this was later confirmed by immunohistochemistry. This is the first report of an outbreak of VER in farmed cod in Norway, and the first report that VER affect cod exceeding 5 g in size.

The Manila clam Ruditapes philippinarum was introduced to Norway in 1987 and was produced in 2 hatcheries until 1991. Clam seed was planted at 6 sites. Two sites were on the Island of Tysnes, south of Bergen. Surviving adult Manila clams were recovered in 1995 and 1996. In the present study, Manila clams from the original seeding that displayed morphological signs of brown ring disease (BRD) were recovered in June 2003 (n = 7) and in June 2004 (n = 17). Samples from extrapallial fluid, tissues and haemolymph were inoculated on marine agar. Replicate subcultures on selective media were used to select potential Vibrio tapetis strains, and in total, 190 bacterial strains were isolated. One of these strains clustered within the V. tapetis clade and was named NRP 45. DNA:DNA hybridisation with the type strain CECT4600 showed 52.7 and 57.3% DNA:DNA similarity. Hybridisation of NRP 45 and the V. tapetis LP2 strain, isolated from corkwing wrasse Symphodus melops, produced 46.6 and 44.4% re-association. Partial gene segments encoding 16S rRNA, gyrase B protein (GyrB) and chaperonin 60 protein (Cpn60) were characterised and compared to CECT 4600. NRP 45 showed 5 differences in the 1416 nucleotides (nt) of the 16S rRNA encoding gene (99.6% similarity), while the GyrB encoding gene had 62 substitutions of 1181 nt compared (94.8% similarity) and the Cpn60 encoding gene had 22 substitutions out of 548 nt compared (96% similarity). This is the first finding of BRD and the first isolation of a V. tapetis-like bacterial strain from a bivalve in Norway. KEY WORDS: Brown ring disease · Vibrio tapetis · Bivalve · Manila clams · Bacteria · NorwayResale or republication not permitted without written consent of the publisher Dis Aquat Org 81: [153][154][155][156][157][158][159][160][161] 2008 disease has subsequently spread along the East Atlantic coast from England to North Africa (Paillard 2004). BRD was first observed in Spain in 1994, in England in 1997 and in Ireland in 1998 (Paillard & Maes 1994, Castro et al. 1996, Novoa et al. 1998. The highest prevalence is encountered in the northernmost countries, and BRD is generally classified as a coldwater disease (Paillard et al. 1994). In Norway, the Manila clam was introduced in 1987 (Mortensen & Strand 2000). Approximately 500 broodstock specimens were imported from Scottish Seafarms Ltd to 2 Norwegian hatcheries, located in Espevik on the Island of Tysnes, south of Bergen, and in Vallersund in South Trøndelag County (Fig. 1A). From 1987 to 1991, more than 200 million seed were produced, with ongrowth done in nurseries at Espevik, Rong, Vallersund and Vågstanda. Most of the Manila clam seed were exported to Spain and Ireland. In Espevik, the native clam R. decussatus was also produced by the same methods and during the same period as the Manila clams. Clam production was terminated in 1991. Between 1988 and 1991, Manila clam seed were released on beaches at 6 sites. At Seløy (Fig.1B), approximately 2000 Manila clam spat were released in 1988, and a total of 28 speci...