Viral Haemorrhagic Septicemia Virus (VHSV) Isolated from Atlantic Herring, Clupea harengus, Causes Mortality in Bath Challenge on Juvenile Herring.

Viral hemorrhagic septicaemia virus (VHSV) has been demonstrated to cause high mortalities in a wide range of teleosts, farmed as well as wild. In Europe, VHSV of genotypes Ib, Id, II, and III have been detected in wild fish, including Atlantic herring Clupea harengus, but disease outbreaks have not been observed in Atlantic herring and the effects on wild stocks are not well documented. Here, we have tested two VHSV isolates from herring (genotypes Ib and III, from the western coasts of Norway and Denmark, respectively) in a challenge experiment with herring (mean weight 2.59 g, SD 0.71 g) caught on the west coast of Denmark. The Norwegian genotype Ib isolate (NO-F-CH/2009) showed an accumulated mortality of 47% compared to 6% mortality with the Danish genotype III isolate 4p168 and zero in the unchallenged control group. In both groups, we found positive rt-RT-PCR and positive immunohistochemistry of VHSV from days 6 and 8 onward. With both isolates, the organs mainly affected were the heart and kidney. The results demonstrate the susceptibility of Atlantic herring to VHSV, and both genotypes gave pathological findings in several organs. Genotype III showed a low mortality rate, and the importance of this genotype for herring is therefore not determined. Genotype Ib showed both high prevalence and mortality, and this genotype is therefore likely to have a negative effect on wild Atlantic herring stocks. Further examinations to determine how VHSV can affect wild Atlantic herring stocks are needed.

Transcriptional regulation of cytokines in the intestine of Atlantic cod fed yeast derived mannan oligosaccharide or ß-glucan and challenged with Vibrio anguillarum.

Immunomodulatory feed additives are expected to exert their primary influence at the intestinal level through the expression of cytokines, which in turn affect the immune responses in fish. In two separate experiments a yeast-derived mannan oligosaccharide product (YM) or a purified ß-glucan (BG) product were fed to Atlantic cod (Gadus morhua L.) for 5 weeks, after which they were bath-challenged with a bacterial pathogen--Vibrio anguillarum. The transcription of selected cytokines (proinflammatory--il1b, il8, ifng; anti-inflammatory--il10) in different intestinal segments was analysed using qPCR. In the case of YM study, the effect of the compound was observed in both the posterior intestine and rectum of Atlantic cod, upon challenge with the pathogen. iIl1b expression in the posterior intestine and rectum of post-challenge fish was significantly higher than that of pre-challenge fish. In the case of il8 the difference was confined to rectum. The expression of ifng was altered only in the anterior intestine upon YM feeding. In the BG trial, the additive had a differential effect on the expression of the cytokine genes. In anterior intestine and rectum, the purified ß-glucan additive significantly elevated the expression of il1b when challenged with V. anguillarum. An effect of BG on the anti-inflammatory cytokine il10 was visible in the rectum after the pathogen challenge. The differential responses of cytokines in the intestine of fish upon exposure to V. anguillarum suggest that both mannan oligosaccharides and ß-glucans impact the ability of Atlantic cod to respond to the pathogen.

Changes in the intestinal microbiota of wild Atlantic cod Gadus morhua L. upon captive rearing.

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.

Susceptibility and Pathology in Juvenile Atlantic Cod Gadus morhua to a Marine Viral Haemorrhagic Septicaemia Virus Isolated from Diseased Rainbow Trout Oncorhynchus mykiss.

The first known outbreak caused by a viral haemorrhagic septicaemia virus (VHSV) strain of genotype III in rainbow trout occurred in 2007 at a marine farm in Storfjorden, Norway. The source of the virus is unknown, and cod and other marine fish around the farms are suspected as a possible reservoir. The main objective of this study was to test the susceptibility of juvenile Atlantic cod to the VHSV isolate from Storfjorden. As the pathology of VHS in cod is sparsely described, an additional aim of the study was to give a histopathological description of the disease. Two separate challenge experiments were carried out, using both intra peritoneal (ip) injection and cohabitation as challenge methods. Mortality in the ip injection experiment leveled at approximately 50% three weeks post challenge. Both immunohistochemical and rRT-PCR analysis of organs sampled from diseased and surviving fish confirmed VHSV infection. No VHSV was detected in the cohabitants. The results indicate that Atlantic cod has a low natural susceptibility to this VHSV genotype III strain. One of the most extensive pathological changes was degeneration of cardiac myocytes. Immunohistochemistry confirmed that the lesions were related to VHSV. In some fish, the hematopoietic tissue of spleen and kidney showed degeneration and immunostaining, classical signs of VHS, as described in rainbow trout. Positive immunostaining of the capillaries of the gills, suggests this organ as a useful alternative when screening for VHSV.

Comparative susceptibility of turbot, halibut, and cod yolk-sac larvae to challenge with Vibrio spp.

In intensive aquaculture systems, high mortalities are frequently observed during the early life stages of marine fish. The aim of this study was to investigate differences in the susceptibility of turbot Scophthalmus maximus, halibut Hippoglossus hippoglossus and cod Gadus morhua to various strains of Vibrio anguillarum (serotypes O1, O2alpha and O2beta), V salmonicida and V splendidus. The bath challenge experiments were performed using a multidish system, with 1 egg well-1. Unchallenged eggs and larvae were used as controls. Larvae in challenged groups that suffered high mortality rates were examined by immunohistochemistry. The overall results with respect to mortality showed that the O2alpha serotype was pathogenic to all 3 species, while the O1 serotype was pathogenic to halibut and cod. The immunohistochemical examinations revealed differences in histopathology. The O1 serotype produced more severe and highly developed infections than the O2alpha serotype. In larvae exposed to the O1 serotype, necrosis and bacterial cells were seen in the dermis, gastrointestinal tract, brain and eye area, while in larvae exposed to the O2alpha serotype, bacteria were usually limited to the gastrointestinal tract. These results suggest either that there are undetermined species differences in host immunity or that these pathogens are host-specific even in the early life stages of fish. The O2beta strain did not cause an increased mortality to halibut and turbot.

Ontogeny of lymphoid organs and development of IgM-bearing cells in Atlantic halibut (Hippoglossus hippoglossus L.).

In teleost fish, the head kidney, thymus, and spleen are generally regarded as important immune organs. In this study, the ontogeny of these organs was studied in Atlantic halibut (Hippoglossus hippoglossus), larvae at various stages of development. We observed that the kidney was present at hatching, the thymus at 33days post hatch (dph), while the spleen was the last organ to be detected at 49dph. All three lymphoid organs were morphologically well developed during late metamorphic stages. Real time RT-PCR analysis showed that IgM mRNA expression could be observed at 66dph and later, which correlates well with in situ hybridisation data showing that a few IgM positive cells could be detected in the anterior kidney and spleen from 66dph. Our data also showed that the highest levels of IgM mRNA could be detected in halibut spleen. Immunostaining using a monoclonal antibody against halibut IgM detected IgM positive cells at 94dph in both the head kidney and the spleen, which is much later than the IgM mRNA. Numerous cells expressing both IgM mRNA and protein could be detected in the spleen and anterior kidney and also to some extent in thymus specimens from adult halibut.

Screening and characterisation of potentially pathogenic bacteria associated with Atlantic cod Gadus morhua larvae: bath challenge trials using a multidish system.

In intensive aquaculture systems, high concentrations of nutrients and high densities of fish larvae provide favorable conditions for opportunistic pathogenic bacteria to flourish. We screened potentially pathogenic bacterial strains isolated from moribund Atlantic cod Gadus morhua larvae, pollack Pollachius pollachius, coalfish Pollachius virens, Atlantic halibut Hippoglossus hippoglossus, rotifers, algae and water samples from different hatcheries. Three identical challenge experiments tested a total of 53 strains. A multidish system was used: cod eggs were placed in single wells, together with 2 ml of sterile seawater, and exposed to the bacterial cultures. Final bacterial concentrations in the wells were 10(6) and 10(4) CFU ml(-1). Eggs and larvae not exposed to bacteria were used as unchallenged controls. Challenged controls were exposed to Vibrio anguillarum strain 610. Eggs were challenged approximately 48 h prior to hatching and mortality was recorded daily throughout the yolk-sac period. In spite of the high challenge dose of 106 CFU ml(-1), only 5 bacterial strains tested caused higher mortality than the unchallenged controls. Four of these strains were identified by 16S rDNA and gyrase B gene (GyrB) sequencing as resembling V. anguillarum and 1 strain resembled Carnobacterium sp. Most of the larvae exposed to these strains died within 10 d of challenge. Serotyping of the strains resembling V. anguillarum gave inconclusive results. This indicates differences in serology compared to the serotypes O1, O2 and O3, associated with disease. Three bacterial strains seemed to have a slower infection rate, indicating a longer incubation period. The remaining 45 strains did not seem to have a negative effect on larval survival, suggesting that these are not primary pathogens.

Immunohistochemistry of Atlantic cod larvae Gadus morhua experimentally challenged with Vibrio anguillarum.

Farming of Atlantic cod Gadus morhua is one of the most rapidly growing sectors of Norwegian aquaculture. Classical vibriosis caused by Vibrio anguillarum is a problem in cod aquaculture. To prevent disease outbreaks, a thorough understanding of the infection route and the impact of the bacteria on the host is important. The intestinal tract, skin and gills have all been proposed as routes of entry for bacterial infections such as vibriosis. We aimed to further develop understanding of V anguillarum serotype O2alpha infections in cod larvae by elucidation of a possible route of entry, the pattern of infection and its histopathology. Cod eggs were transferred to a 24-well polystyrene multi-dish with 2 ml of sterile aerated 80% (28 per thousand salinity) seawater. Challenge doses were 10(4) and 10(6) CFU ml(-1). Unchallenged larvae were used as controls. Larvae for immunohistochemical examination were sampled daily from each group. In most of the larvae, either no or very few bacteria were observed. Typical findings were clusters of bacteria in the spaces between the primary gill lamellae. None of these bacteria seemed to have adhered to the gills. Intestines of 3 out of 161 larvae examined contained positively immunostained bacteria. Some bacteria appeared attached to the microvilli, but none was observed inside epithelial cells. Only 2 larvae from the low-challenge dose group showed clear signs of histopathology, which occurred in the intestine. It is not possible to draw any conclusions regarding the portal of entry.

Vibrio tapetis-like strain isolated from introduced Manila clams Ruditapes philippinarum showing symptoms of brown ring disease in Norway.

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.

The dual myths of the healthy wild fish and the unhealthy farmed fish.

Although diseases, suffering and death have always been recognized as intrinsic parts of life as far as humans are concerned, it seems that many people tend to disregard these factors when it comes to animals. In particular, wild fish are generally assumed to be 'healthy', although the public concept of that term is unclear. In contrast, farmed fish are often popularly viewed as 'unhealthy'. Present knowledge of the importance of epizootics among wild fish is clearly limited, especially regarding viral and bacterial diseases. In contrast to the popular view, the available data indicates that disease among wild fish is common, that epizootics may be of significant ecological importance, and that there is reason to believe that fish diseases among wild as well as cultured fish may be associated with reduced welfare. Large-scale aquaculture without prophylaxis is practically impossible without an unacceptable impact on the environment, as well as reduced fish welfare. In this essay, I oppose the traditional view that industrialisation of aquaculture is linked to reduced fish welfare. In contrast, modern industrial aquaculture with state-of-the-art prophylaxis probably represents a major improvement in controlling fish diseases, thus increasing fish welfare. This is true especially when compared to traditional third world aquaculture, as measured in terms of fish mortality and consumption of antibacterial agents. However, aquaculture may influence diseases of wild fish populations either by providing vectors for transmission of pathogens into new geographic areas, or by altering the balance in host-parasite dynamics by increasing the number of available hosts.

Nodavirus in farmed Atlantic cod Gadus morhua in Norway.

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.

Immunohistochemistry of great scallop Pecten maximus larvae experimentally challenged with pathogenic bacteria.

Three challenge experiments were carried out on larvae of the great scallop Pecten maximus. Larvae were bath-challenged with Vibrio pectenicida and 5 strains resembling Vibrio splendidus and one Pseudoalteromonas sp. Unchallenged larvae were used as negative controls. The challenge protocol was based on the use of a multidish system, where the scallop larvae (10, 13 and 15 d post-hatching in the 3 experiments, respectively) were distributed to 2 ml wells with stagnant seawater and exposed to the bacterial cultures by bath challenge. Presence of the challenge bacteria in the wells was verified by polymerase chain reaction (PCR). A significantly increased mortality was found between 24 and 48 h in most groups challenged with V. pectenicida or V. splendidus-like strains. The exception was found in larval groups challenged with a Pseudoalteromonas sp. LT 13, in which the mortality rate fell in 2 of the 3 challenge experiments. Larvae from the challenge experiments were studied by immunohistochemistry protocol. Examinations of larval groups challenged with V. pectenicida revealed no bacterial cells, despite a high degree of positive immunostaining. In contrast, intact bacterial cells were found in larvae challenged with V. splendidus. In the case of larvae challenged with the Pseudoalteromonas sp., positive immuno-staining was limited to visible bacteria inside the digestive area and cells of the mucosa. The experiments confirm that V. splendidus and V. pectenicida are pathogenic to scallop larvae, and that the Pseudoalteromonas strain is probably not a primary pathogen, although it cannot be ruled out as a secondary pathogen.

Viral and bacterial diseases of Atlantic cod Gadus morhua, their prophylaxis and treatment: a review.

This review summarises the state of knowledge of both viral and bacterial diseases of Atlantic cod Gadus morhua, and their diagnosis, prophylaxis and treatment. The most important losses have been at the larval and juvenile stages, and vibriosis has long been the most important bacterial disease in cod, with Listonella (Vibrio) anguillarum dominant among pathogenic isolates. Vaccination of cod against pathogens such as L. anguillarum and Aeromonas salmonicida clearly demonstrates that the cod immune system possesses an effective memory and appropriate mechanisms sufficient for protection, at least against some diseases. Well-known viruses such as the nodavirus that causes viral encephalopathy and retinopathy (VER), infectious pancreatic necrosis virus (IPNV) and viral haemorrhagic septicaemia virus (VHSV) have been isolated from Atlantic cod and can be a potential problem under intensive rearing conditions. No commercial vaccines against nodavirus are currently available, whereas vaccines against IPNV infections based upon inactivated virus as well as IPNV recombinant antigens are available. A number of investigations of the pharmacokinetic properties of antibacterial agents in cod and their efficacy in treating bacterial infections have been reviewed.

Efficacy of orally administered flumequine in the treatment of vibriosis caused by Listonella anguillarum in atlantic cod Gadus morhua.

The efficacy of orally administered flumequine in the treatment of experimentally induced vibriosis in Atlantic cod Gadus morhua was investigated. Cod (mean +/- SD, 120 +/- 30 g) were randomly distributed to twelve tanks and bath challenged for 1 h with Listonella anguillarum serotype O2alpha, strain HI-610, using a dose of 9.2 x 10(6) CFU ml(-1). At 3 d post-challenge, medication was introduced in 10 of the groups at doses of 2.5, 5, 10, 15 and 25 mg flumequine kg(-1) body weight d(-1) in duplicate. The medication was administered on Days 1, 2, 4, 6, 8, and 10 after the initiation of treatment. In challenged unmedicated fish, mortality started on Day 4 post-challenge, reaching a final cumulative mortality of 82% at Day 18. In the medicated groups, mortality started on Days 3 to 5 post-challenge, reaching final cumulative mortalities of 42, 49, 37, 37 and 23% respectively for the fish treated with 2.5, 5, 10, 15 and 25 mg flumequine kg(-1) body weight d(-1). Survival of medicated fish in all groups was significantly greater than in challenged unmedicated fish (p < 0.001). Twenty-four h following the final medication, HPLC analysis found a linear relationship between doses and mean concentrations of the drug in plasma, muscle and liver.

One-step liquid chromatographic method for the determination of oxytetracycline in fish muscle.

A one-step simple and rapid high performance liquid chromatography (HPLC) method was developed for the determination of oxytetracycline (OTC) in fish tissue. The method involves liquid extraction of muscle tissue, precipitation of proteins and reversed phase HPLC analysis with spectrophotometric detection. The limit of quantitation of OTC in spiked fish muscle was 0.04 microg/g and the method showed high linearity (r(2) = >0.999) in the working range of 0.04-2 microg/g. The precision (%R.S.D.) was between 1.9 and 7.5% for the concentration range 0.04-1.0 microg/g and there was no significant difference between the concentrations determined on three different test days for all four spiked concentrations. The percentage recovery over the spiked concentration range 0.04-1.0 microg/g was consistently within a narrow range of 33-35%. While the method had the advantage of high precision, sensitivity and linearity, the method's additional salient advantages included high sample through-put (60 individual preparations per day) and minimum amount of consumables, time and labour required to perform the analysis. The method was successfully applied to a pharmacokinetic study.

Phylogenetic analysis of bacterial communities associated with larvae of the Atlantic halibut propose succession from a uniform normal flora.

Halibut, the largest of all flatfishes is a valuable species with a great potential for aquaculture. Bacteria play an important role in regulating the health of the early life stages. The present article is the first broad-range molecular analysis of bacterial communities in larvae of the Atlantic halibut (Hippoglossus hippoglossus). DNA was extracted from larvae, water and silo biofilm from hatcheries in Norway, Scotland, Iceland and Canada. Eubacterial 16S rRNA gene fragments were amplified by polymerase chain reaction (PCR) with broad-range primers. Sequences spanning the hyper variable V3 region representing individual bacterial species were separated into community profiles by denaturing gradient gel electrophoresis (DGGE). The profiles revealed simple communities after hatching and bacterial succession following growth. Sequencing and phylogenetic analysis of excised DGGE bands suggested aerobic heterotrophs related to groups of Pseudomonas, Janthinobacterium and possibly Marinomonas to be the primary colonisers of the larvae. After onset of feeding, fermentative species (Vibrio) were detected as well. Comparative analysis of bacterial communities from different geographical regions indicated that larvae of the Atlantic halibut possess a distinct and specific normal flora.

Characterisation of the bacterial community associated with early stages of Great Scallop (Pecten maximus), using denaturing gradient gel electrophoresis (DGGE).

Denaturing gradient gel electrophoresis (DGGE) of PCR-amplified 16S rDNA was used to characterise and compare bacterial communities associated with scallop larvae (Pecten maximus), in different production units in a shellfish hatchery. Water and larvae samples were collected from three different aquaculture systems; stagnant, flow-through and a flow- through system with seawater treated with ozone. Samples were also collected from different algal cultures, inlet tanks and water pipes leading to the different aquaculture systems. Clear differences were seen between the bacterial community associated with the larvae and in the water from the different aquaculture systems. However, there were high similarities in the community composition between different water samples and between larvae samples collected at different time periods, indicating a high stability in the bacterial communities. Fifty three percent of the sequences from these samples were similar to 16S rRNA gene sequences of members of the gamma-subclass of the Proteobacteria. The different algal cultures had different bacterial communities, however 73 percent of the sequences were similar to 16S rRNA gene sequences of members of the alpha-subclass of the Proteobacteria. Differences in the DGGE profiles were also seen between the samples taken from the inlet tanks and water pipes, indicating a change in the bacterial community composition as the water passed through the pipes. To our knowledge this is the first study investigating bacterial communities associated with Great Scallop larvae in different aquaculture systems including noncultured components.

Selection and identification of autochthonous potential probiotic bacteria from turbot larvae (Scophthalmus maximus) rearing units.

The purpose of this study was to select, identify and characterise bacteria as a disease control measure in the rearing of marine fish larvae (turbot, Scophthalmus maximus). Thirty-four out of 400 marine bacterial strains exhibited in vitro anti-bacterial activity against three fish larval pathogens. Two strains originated from culture collections and thirty two strains were isolated directly from turbot larvae rearing units using a pre-selection procedure to facilitate detection of antagonists. Approximately 8,500 colonies from colony-count plates were replica-plated on agar seeded with Vibrio anguillarum, and 196 of them caused zones of clearing in the V. anguillarum agar layer. Of these, 32 strains exhibited reproducible antibacterial properties in vitro when tested against the fish pathogens V. anguillarum 90-11-287, V. splendidus DMC-1 and a Pseudoalteromonas HQ. Seventeen antagonists were identified as Vibrio spp. and four of twelve tested were lethal to yolk-sac larvae. The 15 remaining strains were identified as Roseobacter spp. based on phenotypic criteria and 16S rDNA gene sequence analysis of two strains representing the two major RAPD groups. Most of the remaining 164 strains selected in the initial replica plating were identified as Vibrionaceae or Pseudoalteromonas. Roseobacter spp. were not lethal to egg yolk sac turbot larvae and in two of three trials, the mortality of larvae decreased (p > 0.001) in treatments where 10(7) cfu/ml Roseobacter sp. strain 27-4 was added, indicating a probiotic potential.

Pharmacokinetics of florfenicol in cod Gadus morhua and in vitro antibacterial activity against Vibrio anguillarum.

The pharmacokinetic profile of the antibacterial agent florfenicol was studied in plasma after intravenous (i.v.) injection and in plasma, muscle and liver following oral (p.o.) administration to cod Gadus morhua, held in seawater at 8 degrees C and weighing 100 to 200 g. Following i.v. injection, the plasma drug concentration-time profile showed 2 distinct phases. The plasma distribution half-life (t1/2alpha) was estimated to be 1.6 h, the elimination half-life (t1/2beta) to be 43 h, the total body clearance (ClT) to be 0.015 1 kg(-1) h(-1) and mean residence time (MRT) to be 74 h. The volume of distribution at steady state, Vd(ss), was calculated to be 1.1 l kg(-1). Following p.o. administration, the bioavailability was estimated to be 91%, the peak plasma concentrations (Cmax) to be 10.8 microg ml(-1) and the time to peak plasma concentrations (Tmax) to be 7 h. Corresponding Cmax and Tmax values were 13.0 microg g(-1) and 9 h, respectively, in muscle and 12.1 microg g(-1) and 9 h, respectively, in liver. The in vitro minimum inhibitory concentration (MIC) values of florfenicol against 3 Vibrio anguillarum strains isolated from diseased cod (A-21, HI-610, HI-618) were 0.5 microg ml(-1) for all 3 strains.

Characterization of strains of Vibrio splendidus and V. tapetis isolated from corkwing wrasse Symphodus melops suffering vibriosis.

Two vibrio bacteria pathogenic to the corkwing wrasse Symphodus melops were isolated. Vibriosis-inducing strain LP1 was isolated as the dominanting bacterium in kidney samples of dead and moribund wrasse from a population suffering vibriosis and high daily mortality in 1998 on the Norwegian west coast. The other vibriosis-inducing strain, LP2, was isolated from wrasse captured the following year. Re-infection experiments have confirmed that these strains cause vibriosis in corkwing wrasse. Both strains were typical vibrios sharing the traits of fermentative Gram-negative curved rods with motility and a positive oxidase reaction. Detailed biochemical and genetic characterisation revealed a close affiliation to known species of the marine environment. The first isolate, LP1, is a form of the ubiquitous seawater organism Vibrio splendidus, while the second isolate, LP2, is closely related to V. tapetis (previously only known as the brown ring disease agent in clams). Identification of the new wrasse pathogens V. splendidus LP1 and V. tapetis LP2 is facilitated by break points observed in this study.