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Lymphocystis disease is a geographically widespread disease affecting more than 150 different species of marine and freshwater fish. The disease, provoked by the iridovirus lymphocystis disease virus (LCDV), is characterized by the appearance of papillomalike lesions on the skin of affected animals that usually self-resolve over time. Development of the disease is usually associated with several environmental factors and, more frequently, with stress conditions provoked by the intensive culture conditions present in fish farms. In gilthead sea bream (Sparus aurata), an economically important cultured fish species in the Mediterranean area, a distinct LCDV has been identified but not yet completely characterized. We have used direct sequencing of the virome of lymphocystis lesions from affected S. aurata fish to obtain the complete genome of a new LCDV-Sa species that is the largest vertebrate iridovirus sequenced to date. Importantly, this approach allowed us to assemble the full-length circular genome sequence of two previously unknown viruses belonging to the papillomaviruses and polyomaviruses, termed Sparus aurata papillomavirus 1 (SaPV1) and Sparus aurata polyomavirus 1 (SaPyV1), respectively. Epidemiological surveys showed that lymphocystis disease was frequently associated with the concurrent appearance of one or both of the new viruses. SaPV1 has unique characteristics, such as an intron within the L1 gene, and as the first member of the Papillomaviridae family described in fish, provides evidence for a more ancient origin of this family than previously thought. IMPORTANCELymphocystis disease affects marine and freshwater fish species worldwide. It is characterized by the appearance of papillomalike lesions on the skin that contain heavily enlarged cells (lymphocysts). The causative agent is the lymphocystis disease virus (LCDV), a large icosahedral virus of the family Iridoviridae. In the Mediterranean area, the gilthead sea bream (Sparus aurata), an important farmed fish, is frequently affected. Using next-generation sequencing, we have identified within S. aurata lymphocystis lesions the concurrent presence of an additional LCDV species (LCDV-Sa) as well as two novel viruses. These are members of polyomavirus and papillomavirus families, and here we report them to be frequently associated with the presence of lymphocysts in affected fish. Because papillomaviruses have not been described in fish before, these findings support a more ancient origin of this virus family than previously thought and evolutionary implications are discussed. L ymphocystis is a widely distributed disease affecting over 150 different marine and freshwater fish species (1). It was initially described during the 19th century and is characterized by the appearance of papillomalike lesions on the skin and fins, which develop over prolonged periods of time ranging from weeks to months. Occasionally, lesions on internal organs have been described, but the condition is rarely life-threatening and the outgrowths usually self-r...

The in vivo and in vitro toxicity of bacterial cells and their extracellular products (ECPs) from 16 strains of Photobacterium damselae subsp. damselae isolated from 7 epizootic outbreaks were evaluated. On the basis of their 50% lethal dose (LD 50 ) values (about 1 × 10 5 CFU), these strains may be considered as moderately virulent. However, their ECPs were strongly lethal for redbanded seabream Pagrus auriga causing fish death within 2 h post-inoculation (protein concentration ranged between 2.1 and 6.41 µg g -1 fish). The bacterial ECPs tested exhibited several enzymatic activities, such as amylase, lipase, phospholipase, alkaline phosphatase, esterase-lipase, acid phosphatase, and β-glucosaminidase. These ECPs displayed a strong cytotoxic effect on 4 fish and 2 mammalian cell lines, although this activity disappeared when ECPs were heated at 100°C. The virulence of the strains tested could not be related to the hemolytic activity or to the production of the toxin damselysin. Therefore, another unknown type of toxin could play an important role in the virulence mechanisms of this bacterial pathogen.KEY WORDS: Toxicity · ECP · Photobacterium damselae subsp. damselae · Cultured marine fish Resale or republication not permitted without written consent of the publisherDis Aquat Org 92: [31][32][33][34][35][36][37][38][39][40] 2010 brane, has been described (Kreger 1984, Kothary & Kreger 1985, Kreger et al. 1987. In addition, a relationship between the degree of virulence and the hemolytic activity has been demonstrated in P. damselae subsp. damselae strains isolated from fish (Fouz et al. 1993). However, Cutter & Kreger (1990) found that not all the P. damselae subsp. damselae strains presented the damselysin gene (dly), but only those strains showing intense hemolytic activity. A further study demonstrated that the presence of this gene was not correlated to the virulence in mice and fish of 17 P. damselae subsp. damselae strains isolated from different sources (Osorio et al. 2000).The extracellular products (ECPs) are produced by bacterial pathogens to facilitate the uptake of nutrients from the surrounding environment, and/or for the successful penetration and survival of pathogens inside the host (Bakopoulos et al. 2003). However, the role of ECPs in the pathogenesis of Photobacterium damselae subsp. damselae in fish is poorly known, which is a considerable disadvantage for the development of vaccines and vaccine strategies, since it has been suggested that the ECP components are major antigenic compounds of several vaccine formulations (Collado et al. 2000, Bakopoulos et al. 2004.The aim of this study was to determine the toxicity of different Photobacterium damselae subsp. damselae strains isolated from cultured diseased fish. For this purpose, we performed in vivo and in vitro assays using bacterial cultures and their ECPs. In addition, the enzymatic activities of the ECPs and their cytotoxicity in fish and mammalian cell lines were compared. MATERIALS AND METHODSBacterial strains. In this...

The aim of this study was to determine the in£uence of the feeding regimes in Senegalese sole (Solea senegalensis) cultured under extensive, semi-extensive and intensive production systems. A total of 254 bacterial isolates from guts of ¢sh cultured under di¡erent production systems and feeding regimes were tested. Biochemical tests and genetic analyses based on the 16S rDNA sequence analysis were conduced to identify bacterial strains.Vibrio species were the most represented taxonomic group in the culturable microbiota of S. senegalensis guts tested. Particularly, Vibrio ichthyoenteri was the most frequently isolatedVibrio species. Comparison among diets showed a signi¢cant reduction (Po0.05) in vibrio percentages and a higher occurrence of Shewanella species in Senegalese soles fed polychaeta. In addition, a major in£uence of environmental temperature on microbiota composition was detected. Cold temperatures brought about a change in the percentages of Vibrio species and a higher representation of a-Proteobacteria in both outdoor systems (extensive and semi-extensive). The signi¢cant di¡erences between intestinal bacterial composition in Senegalese soles fed commercial diets and natural preys (polychaeta) reveal the necessity to develop speci¢c optimized diets for the intensive rearing of this ¢sh species.

Nanocomposites and hybrid materials of Ag−1,3,5benzenetricarboxylic acid metal−organic frameworks (MOFs) with Sand N-carbon quantum dots (CQDs) were synthesized and evaluated for their antibacterial activity against representative Gram-positive (Bacillus subtilis) and Gram-negative (Escherichia coli) bacterial strains using the qualitative disk-diffusion approach and the quantitative minimum inhibitory concentration test. The composites and hybrids were found to be nontoxic to living cells. The composite formation fostered a synergistic effect that enhanced their antibacterial activity compared with those of their pristine components. Charge transfer from AgMOF to CQDs facilitated the electrostatic interactions of the composites and hybrids with the bacterial cell membranes. Enhanced bactericidal activity was linked to morphological features (a nanorod-like morphology) and specific surface chemistry. The latter affected the release of silver. Silver on the surface of the MOFs rather than silver in the bulk was found to be important. The destruction of the MOF component in the extracellular environment led to the release of silver ions, which have a high affinity to S compounds of the cell physiology. The formation of metallic silver (Ag°) and silver sulfides (Ag 2 S) was suggested as essential for the ability of the composites and hybrids to inhibit bacterial growth. To the best of our knowledge, this is the first study that introduces the bactericidal effect of AgMOF−CQDs composites and hybrids.

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This report describes a viral epidemiological study of wild fish around the Gulf of Cadiz (southwestern Iberian Peninsula) and is focused on infectious pancreatic necrosis virus (IPNV), viral hemorrhagic septicemia virus (VHSV), and viral nervous necrosis virus (VNNV). One fish species (Chelon labrosus) was sampled inside the gulf, at the mouth of the San Pedro River. Another 29 were sampled, in three oceanographic campaigns, at sites around the Bay of Cadiz. The fish were processed individually and subjected to isolation in cell culture and molecular diagnosis. VHSV was not isolated from any species. Thirteen IPNV-type isolates were obtained from barracuda (Sphyraena sphyraena), axillary seabream (Pagellus acarne), common two-banded seabream (Diplodus vulgaris), common pandora (P. erythrinus), Senegal seabream (D. bellottii), and surmullet (Mullus surmuletus). Six VNNV isolates were obtained from axillary seabream, common pandora, black seabream (Spondyliosoma cantharus), red mullet (Mullet barbatus), Lusitanian toadfish (Halobatrachus didactylus), and tub gurnard (Chelidonichtys lucerna). In the river mouth, viruses were detected only after reamplification, obtaining prevalence percentages of IPNV and VNNV (44.4 and 63.0%, respectively) much higher than those observed in the oceanographic campaigns (25.7 and 19.6%, respectively). The opposite results were obtained in the case of VHSV after reamplification: 11.1% in the river mouth and 43.6% in the oceanic locations. Analyzing the results with respect to the proximity of the sampling sites to the coast, an anthropogenic influence on wild fish is suggested and discussed. The type of viruses and the presence of natural reassortants are also discussed. Microbial pathogens and potential hosts have coexisted in the marine environment for a long time. This coexistence allows the achievement of a balance between the pathogen and its host: The pathogen self-limits its virulence to reduce the negative effect on specific hosts, and the host uses defense strategies to minimize the negative consequence of the infection. The stressful conditions of intensive culture, however, usually break that equilibrium, and consequently, disease episodes probably occur more frequently than in natural environments.Among pathogens, the relevance of viral fish diseases relies on the high morbidity and mortality rates of infections and on the lack of effective treatments and prophylactic measures. In addition, several factors such as the vertical transmission of some fish viruses and the ability to establish chronic infections or carrier states among survivors must also be considered (1).The Ninth Report of the International Committee on Taxonomy of Viruses (2) formally recognizes 40 species of fish viruses in 11 taxonomic families. However, only a few studies of wild fish viruses have been performed, in contrast to those done with cultured or ornamental fish. This is probably due to the important losses that viral diseases cause in cultured fish. Moreover, since diseased or moribund animal...