The concentration of bacteriophages in natural unpolluted waters is in general believed to be low, and they have therefore been considered ecologically unimportant. Using a new method for quantitative enumeration, we have found up to 2.5 x 10(8) virus particles per millilitre in natural waters. These concentrations indicate that virus infection may be an important factor in the ecological control of planktonic micro-organisms, and that viruses might mediate genetic exchange among bacteria in natural aquatic environments.

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For most marine aquaculture species, one of the main bottlenecks is the stable production of high quality juveniles. The high and unpredictable mortality in the first weeks after hatching of marine fish larvae remains a challenging problem that needs to be solved. The severity of the problem differs between species, but cannot be considered adequately solved for any species. Both scientific evidence and experience in hatcheries for a variety of fish, shrimp and shellfish species are accumulating as support for the hypothesis that detrimental fish–microbe interactions are the cause of these problems. Host–microbe interactions in reared fish are still poorly understood, except for a few pathogens, and empirical data of the quality required to test this hypothesis, are lacking. This article provides an overview on the current knowledge of the microbial environment of fish larvae, including methodological aspects to characterize the microbial community (both using culture‐dependent and culture‐independent methods). Further, the current knowledge of the immunology of fish larvae is reviewed, including recent advances in the understanding of toll‐like receptors, inflammatory cytokines, mast cells and piscidins, and the ontogeny of the adaptive immune system. Finally, we provide an overview of the state of the art with respect to steering of microbial communities associated with fish larvae – both steering of community composition and of its activity (e.g. by quorum sensing interference).

Phaeobacter gallaeciensis can antagonize fish-pathogenic bacteria in vitro, and the purpose of this study was to evaluate the organism as a probiont for marine fish larvae and their feed cultures. An in vivo mechanism of action of the antagonistic probiotic bacterium is suggested using a non-antagonistic mutant. P. gallaeciensis was readily established in axenic cultures of the two microalgae Tetraselmis suecica and Nannochloropsis oculata, and of the rotifer Brachionus plicatilis. P. gallaeciensis reached densities of 107 cfu/ml and did not adversely affect growth of algae or rotifers. Vibrio anguillarum was significantly reduced by wild-type P. gallaeciensis, when introduced into these cultures. A P. gallaeciensis mutant that did not produce the antibacterial compound tropodithietic acid (TDA) did not reduce V. anguillarum numbers, suggesting that production of the antibacterial compound is important for the antagonistic properties of P. gallaeciensis. The ability of P. gallaeciensis to protect fish larvae from vibriosis was determined in a bath challenge experiment using a multidish system with 1 larva per well. Unchallenged larvae reached 40% accumulated mortality which increased to 100% when infected with V. anguillarum. P. gallaeciensis reduced the mortality of challenged cod larvae (Gadus morhua) to 10%, significantly below the levels of both the challenged and the unchallenged larvae. The TDA mutant reduced mortality of the cod larvae in some of the replicates, although to a much lesser extent than the wild type. It is concluded that P. gallaeciensis is a promising probiont in marine larviculture and that TDA production likely contributes to its probiotic effect.

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33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50The purpose of this study was to evaluate the probiotic effect of the marine bacterium Roseobacter strain 27-4 in turbot larvae infected with the pathogen Vibrio (Listonella) anguillarum. Initial trials demonstrated that cells of Roseobacter were not harmful to larvae whereas, large amounts of bacterial culture supernatant caused rapid mortality (70% at day 10 compared to 20% in the control). A similar high mortality was, however, also seen, when sterile marine broth was added to the larvae. Presumably both types of medium enhanced growth of opportunistic pathogens. In subsequent trials, both a pathogen, Vibrio anguillarum, and the probiont, Roseobacter strain 27-4, were delivered to the larvae bioencapsulated in rotifers. Accumulated mortality of Vibrio infected larvae increased to 80-90% over 10 days, whereas, mortality in non-infected controls was significantly lower (60-70%). Feeding larvae with rotifers enriched with Roseobacter 27-4 parallel to V. anguillarum infection, brought the accumulated mortality to the level of control indicating a clear in vivo effect. Roseobacter 27-4 could be detected in larvae both by agar plating and by immunohistochemistry, being located in the gastrointestinal lumen, and apparently did not colonise the larval gut and intestinal epithelium. Plate counts decreased when enriched feed was no longer added, suggesting that the probiont, Roseobacter 27-4, should be supplied repeatedly to exert its positive effect. Introduction

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.

ABSTRACT. The susceptibility of the Atlantic halibut Hjppoglossus hippoglossus yolk-sac larvae to viral encephalopathy and retinopathy (VER) was investigated by waterborne challenge experiments with nodavuus. Transfer of VER was inhcated by several hnes of evidence. A sigluficantly higher cumulative mortality was observed after challenge with virus compared to mock challenge, and increasing doses of vlrus resulted in shorter incubation periods. When the challenge was performed on the day after hatchmg, the time from inoculation to the time when 50% of the larvae were dead (LT,,) ranged from 26 to 32 d. Postponement of challenge for 13 d reduced the LT,, to 14 d, inhcating that the susceptibility of the larvae to the present nodavirus strain was low during the first 2 wk after hatching. The progression of the infect~on was monitored by sequential unmunohistochemistry and electron mcroscopy. On Day 18 after hatching the initial signs of infection were observed as a prominent focus of immunolabelling in the caudal part of the brain stem. In the same larvae irnrnunolabelled single cell lesions were observed in the stratified epithelium of the cranial part of the intestine. The portal of entry into the larvae may thus have been the intestinal epithelium, while the route of infection to the CNS may have been axonal transport to the brain stem through cranial nerves such as the vagus nerves. Later in the infection, lesions became more severe and widespread and were also found throughout the b r a~n and spinal cord and in the retina, cranial ganglia, intestme, liver, olfactory epithelium, yolk-sac epithelium, gills and pectoral fins. The mortality in all virus-challenged groups was 100%. This study thus demonstrates that the present nodavirus strain is able to replicate and cause VER in Atlantic halibut yolk-sac larvae at temperatures as low as 6°C