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1.
Photobacterium damsela ssp. piscicida (Phdp) isolates were grown in various bacteriological media, in eukaryotic cell culture media and in the presence of fish cells (resembling some aspects of in vivo growth environments). Bacterial cells, extracellular products (ECPs) and crude capsular polysaccharide were isolated and analysed by electrophoresis and Western blot using sea bass sera. Growth in bacteriological media conserved the synthesis of cell and extracellular components when these were compared with those prepared under near-in vivo growth conditions. In fact, synthesis of a larger range of cell components was induced after growth in bacteriological media. Certain media based on yeast extract and peptones from various sources and a specific salt formulation induced the synthesis of novel cell components at approximately 21.3 and 14 kDa. These antigens were recognized by sea bass sera collected after natural pasteurellosis outbreaks and other sea bass sera raised against live or inactivated Phdp cells. The ECPs of the pathogen were not good immunogens in their soluble form despite various treatments prior to immunization. The results are discussed with respect to vaccine development.  相似文献   

2.
ABSTRACT:   Seawater fishes are affected by a pathology commonly called 'myxobacteriosis', caused by Tenacibaculum maritimum (formerly Flexibacter maritimus ). The disease is characterized by fin erosion and necrotic ulcers of skin and muscle, and by low but constant mortality in cultured marine fish; in Italy is one of the most important and widely spread diseases affecting sea bass Dicentrarchus labrax , gilthead seabream Sparus aurata , sharp-snouted bream Diplodus puntazzo , white bream Diplodus sargus , and six-tooted bream Dentex dentex . In order to obtain an effective vaccine against the disease, formalin killed cells (FKC), extracellular products (ECPs) and crude lipopolysaccharide (LPS) preparations were obtained from the T. maritimum strain SPVId and injected intraperitoneally twice into the sea bass. The fish immune response to the preparations was studied: agglutinating antibody titer and in vitro phagocytosis were determined after the first and second injection in order to evaluate whether the preparations are immunogenic or not and if the booster effect took place. The results show that FKC and LPS preparations increased the antibody titer after the first injection when compared to the control sea bass. Moreover, all the preparations stimulated a secondary (booster) response. In vitro phagocytosis of the total blood was significantly higher for all the preparations when compared to the controls, but the crude LPS immunized sea bass showed the highest activity.  相似文献   

3.
Bacterial cells of the marine fish pathogen Photobacterium damsela subsp. piscicida were grown in novel culture media. A mixture of whole cells and extracellular components was inactivated and used in bath, intraperitoneal (i.p.) and oral vaccination of sea bass, Dicentrarchus labrax, employing two sizes of fish. A commercial vaccine was used for comparative purposes. Control and immunized fish were either bath or intraperitoneally challenged 6 and 12 weeks post-vaccination. Small fish had significantly higher relative percentage survival with the novel vaccine mixture both at 6 and 12 weeks post-vaccination by bath, in comparison with the commercial vaccine. No protection was afforded at 6 or 12 weeks post-immunization by either vaccine after challenge via i.p. injection. Sea bass (1.5-2 g) intraperitoneally vaccinated with various adjuvanted vaccine mixtures were not protected against pasteurellosis. In contrast, larger sea bass (20 g) benefited from vaccination with the novel vaccine mixtures. Intraperitoneal challenge with the pathogen resulted in protection in both fish groups vaccinated with novel vaccine mixtures, whereas control fish suffered high mortalities (> 80%). Orally vaccinated fish were immersion challenged with the pathogen. At 6 and 12 weeks post-vaccination the control fish had a high mortality and the fish vaccinated with the novel vaccine mixture achieved good protection.  相似文献   

4.
As adhesion and translocation through fish gut enterocytes of the pathogen Vibrio (Listonella) anguillarum are not well investigated, the effective cause of disease and mortality outbreaks in larval sea bass, Dicentrarchus labrax, suffering from vibriosis is unknown. We detected V. anguillarum within the gut of experimentally infected gnotobiotic sea bass larvae using transmission electron microscopy and immunogold labelling. Intact bacteria were observed in close contact with the apical brush border in the gut lumen. Enterocytes contained lysosomes positive for protein A‐gold particles suggesting intracellular elimination of bacterial fragments. Shed intestinal cells were regularly visualized in the gut lumen in late stages of exposure. Some of the luminal cells showed invagination and putative engulfment of bacterial structures by pseudopod‐like formations. The engulfed structures were positive for protein A‐colloidal gold indicating that these structures were V. anguillarum. Immunogold positive thread‐like structures secreted by V. anguillarum suggested the presence of outer membrane vesicles (MVs) hypothesizing that MVs are potent transporters of active virulence factors to sea bass gut cells suggestive for a substantial role in biofilm formation and pathogenesis. We put forward the hypothesis that MVs are important in the pathogenesis of V. anguillarum in sea bass larvae.  相似文献   

5.
Bacterial subcellular components and probiotics were successful for the stimulation of immunity and the prevention of Vibrio harveyi infections in rainbow trout, Oncorhynchus mykiss (Walbaum). Rainbow trout were immunized with whole inactivated cells of V. harveyi to obtain polyclonal antibodies against specific antigens. Western blotting showed a unique reactive band (∼93 kDa) between serum and bacterial proteins from outer membrane proteins (OMP) and extracellular products (ECP). Probiotics were selected according to their capability to inhibit V. harveyi . Two of these bacteria, i.e. A3-47 and A3-51, showed cross-reactivity with V. harveyi antiserum. Their OMPs and ECPs were reactive with V. harveyi antiserum in bands of ∼93 kDa for A3-51 and higher for A3-47. In vivo tests determined that fish fed with A3-51 produced cross-reactive antibodies against V. harveyi and also, the survival of these fish infected with V. harveyi was high, being similar to the level achieved with vaccinated fish. Thus, the probiotics, when administered as live preparations, were capable of producing cross-reactive antibody against specific bacterial pathogens.  相似文献   

6.
This work was carried out to investigate the effects of injection of Tenacibaculum maritimum formalin‐killed cells (FKC), extracellular products (ECPs) and crude lipopolysaccharide (LPS) as well as 1% feed supplements of oil extracts of Echinacea purpurea and Origanum vulgare on sea bass immunity improvement to the favour of T. maritimum experimental infection control after 4 weeks of the experiment. Tenacibaculum maritimum isolated from naturally infected sea bass showed brown to yellowish‐brown lesions (sores) on gills, skin and/or fins and identified by different biochemical methods and polymerase chain reaction technique. Immune parameters namely, total protein, globulin and lysozyme activity, as well as the relative level of protection were improved by T. maritimum (FKC), (LPS), (ECPs), O. vulgare and E. purpurea, respectively compared with control. Histopathological examination of T. maritimum naturally infected sea bass indicated many pathological changes in gill, skin and musculatures. Present study could be concluded that application of T. maritimum (FKC), (LPS), (ECPs), O. vulgare or E. purpurea improved sea bass immunity to the favour of disease resistance against T. maritimum. Further investigations on the combination between the previous control methods and the vaccine application methods will be needed.  相似文献   

7.
Francisella orientalis is a highly virulent, emerging bacterium that causes mass mortalities in tilapia. This pathogen also affects numerous other warm-water fish species, including three-line grunt, hybrid striped bass and various ornamental fish. This study sheds light on two new species of fish that are susceptible to F. orientalis. Asian seabass and largemouth bass showed variable levels of susceptibility in a bacterial challenge experiment. After intraperitoneally injected with a dose of 106 CFU/fish, a total of 64.28% and 21.42% mortalities were obtained in Asian seabass and largemouth bass, respectively. Meanwhile, Nile tilapia showed acute mortality of 100%. All fish showed typical lesions of francisellosis, including multifocal granulomas in the spleen and head kidney. Immunohistochemical analysis revealed strong positive signals inside the granulomas of all fish. The bacterial recovery in solid media from infected fish was highest in Nile tilapia (85.71%), followed by Asian seabass (35.71%) and largemouth bass (21.42%). PCR results tested 100% positive for Nile tilapia, and 78.57% and 21.42% for Asian seabass and largemouth bass, respectively. In conclusion, Asian seabass and largemouth bass are susceptible to this pathogen, which warrants new management strategies when employing predation polyculture systems of these species with tilapia.  相似文献   

8.
In the Mediterranean area, wild fish have often been suggested as either the reservoirs of the causative agents or at least the carriers of the pathogens responsible for disease outbreaks in cultured fish. However, no epidemiological investigations on actual pathogen/disease interactions between farmed and wild fish have been conducted even for the most important fish pathogens. Only sporadic isolations and identifications of various pathogens in wild fish have been done and real associations with the pathological conditions that exist within the farm environment and vice versa have not been established. Monogenean ectoparasite Sparicotyle chrysophrii and isopod Ceratothoa oestroides are commonly found in the Mediterranean cage-reared sea bream and sea bass and in the surrounding wild fish population. Both species were recognized as pathogens that seasonally inflict serious losses in fingerlings and juveniles of sea bream and sea bass, being potentially propagated and exchanged during wild and caged fish interaction. In order to evaluate the degree of pathogens transfer between wild and farmed fish, we investigated genetic population structures of these two important parasitic pathogens inferred by mtDNA cytochrome oxidase I locus. Parasites isolated from wild and farmed fish on the two most productive Adriatic fish farms showed genetic heterogeneity, contradicting widely accepted hypothesis of cross-contamination, at least in case of S. chrysophrii and C. oestroides. As far as we know, this is the first report that in a study of pathogen transfer molecular evidence was employed to asses the genetic population structure of shared parasites.  相似文献   

9.
The antigenicity of extracellular products (ECPs) derived from Mycobacterium spp. isolated from snakehead, Channa striata (Bloch), and Siamese fighting fish, Betta splendens (Regan), were examined by immunoblotting and enzyme-linked immunosorbent assay (ELISA) using sera collected from immunized rabbits, mice and fish (rainbow trout). All three species responded to a 65-kDa protein present in both the ECPs and whole cell sonicates (WCSs) from a variety of Mycobacterium spp. Cross-reactivity of anti- M. tuberculosis and anti-human heat-shock protein monoclonal antibodies (MAbs), and the presence of fibronectin binding proteins secreted into ECPs of mycobacteria were also examined. The MAbs against human 60-kDa heat-shock protein cross- reacted with the band at 65 kDa in the ECPs of TB1 (isolated from snakehead fish) and the type strain M. marinum, while the anti- M. tuberculosis MAb F29–47 elicited a strong reaction with a band at 21 kDa with most of the ECPs from mycobacterial strains examined. The major fibronectin-binding proteins were located between 21 and 25 kDa. The 65-kDa protein from ECPs of Mycobacterium spp. proved strongly immunogenic to rabbits, mice and fish. Rabbit antiserum against the 65-kDa protein from strain TB267 reacted with many non- Mycobacterium WCSs, and therefore, the 65-kDa protein from Mycobacterium spp. is believed to be a common protein found in many fish bacterial pathogens.  相似文献   

10.
In this study, the proteome profile of European sea bass (Dicentrarchus labrax) muscle was analyzed using two-dimensional electrophoresis (2-DE) and tandem mass spectrometry with the aim of providing a more detailed characterization of its specific protein expression profile. A highly populated and well-resolved 2-DE map of the sea bass muscle tissue was generated, and the corresponding protein identity was provided for a total of 49 abundant protein spots. Upon Ingenuity Pathway Analysis, the proteins mapped in the sea bass muscle profile were mostly related to glycolysis and to the muscle myofibril structure, together with other biological activities crucial to fish muscle metabolism and contraction, and therefore to fish locomotor performance. The data presented in this work provide important and novel information on the sea bass muscle tissue-specific protein expression, which can be useful for future studies aimed to improve seafood traceability, food safety/risk management and authentication analysis. This work is also important for understanding the proteome map of the sea bass toward establishing the animal as a potential model for muscular studies.  相似文献   

11.
Channel catfish, Ictalurus punctatus (Rafinesque), goldfish, Carassius auratus (L.), striped bass, Morone saxatilis (Walbaum), and Gulf sturgeon, Acipenser oxyrinchus desotoi Vladykov, were acclimatized to fresh water or salinities of 9.0‰ or less and then exposed to Flavobacterium columnare (formerly known as Flexibacter columnaris ), the bacterial pathogen that causes columnaris disease. None of the fish acclimatized to 3.0 or 9.0‰ salinity died, and all deaths in lower salinities occurred between 1 and 5 days after exposure to F. columnare . Mortality was 97.7% in fresh water and 67.1% in 1.0‰ salinity for channel catfish (model SE, 1.8) and 66.5% in fresh water and 40.8% in 1.0‰ salinity for goldfish (model SE, 1.2); and 96.9% in fresh water and 61.7% in 1.0‰ salinity for striped bass (model SE, 1.8). After exposure to F. columnare , none of the Gulf sturgeon died. Flavobacterium columnare was isolated from the skin and gills of all fish dying during the experiments, but was not isolated from survivors in fresh water and 1.0‰ salinity 21 days after bacterial exposure. In vitro growth of bacteria was significantly higher in 1.0 or 3.0‰ salinity than in control medium (0.3‰ salinity). However, in vitro adhesion of bacteria was reduced with increasing salinity, which could explain the lower mortality of fish at higher salinities.  相似文献   

12.
The antibacterial effect of Lactococcus lactis subsp. lactis against the fish pathogen Vibrio anguillarum was determined in Artemia franciscana nauplii and in European sea bass (Dicentrarchus labrax) larvae fed with ten pulses of nauplii enriched with L. lactis. The evaluation of the bactericidal activity of the extracellular products of L. lactis in vitro showed inhibition of growth of Vibrio (Listonella) anguillarum and Photobacterium damselae subsp. piscicida. The incorporation of L. lactis in Artemia nauplii did not affect their survival and offered protection in a challenge with V. anguillarum, significantly increasing LD50. The administration of Artemia nauplii enriched with L. lactis for 48 h to sea bass larvae for five consecutive days had no adverse effect on survival of fish. In an in vivo challenge test with V. anguillarum using sea bass larvae, fish treated with nauplii enriched with the probiotic L. lactis showed significantly (P < 0.001) increased survival rates of 81 % compared with the untreated group of challenged fish (24 %). Our results indicate that L. lactis is a probiotic suitable to be used for the prevention of vibriosis in fish larvae and can be safely administered through their live feed Artemia nauplii.  相似文献   

13.
The objective of the present study was to investigate the combined effect of several dietary contents of vitamin E and polyunsaturated fatty acids (PUFA), mainly docosahexaenoic acid (DHA), on growth, survival, biochemical composition and tissue morphology of sea bass along early development. A feeding experiment was conducted in sea bass larvae using five different diets with the same proximate composition and different ratios of DHA concentrated fish oil [10, 30 and 50 g kg?1 dry weight (DW)] and vitamin E (α‐tocopherol acetate) (1500 and 3000 mg kg?1 DW). DHA was readily deposited in fish tissues and associated with higher sea bass mortalities probably because of increased peroxidation risks. Besides, the elevation of dietary DHA contents up to 5% severely increased the incidence of muscular lesions and the presence of ceroid pigment within hepatocytes. However, elevation of dietary vitamin E levels markedly reduced the incidence of these symptoms in sea bass, increasing the tissue content in several PUFA and improving growth and stress resistance. Moreover, when sea bass was fed diets containing high vitamin E levels, fish showed a significant improvement in growth when dietary DHA was raised from 1% to 3%. Therefore, in sea bass larvae, a ratio of 30 g kg?1 DHA and 3000 mg kg?1 vitamin E seems to be adequate to achieve a good larval performance and to avoid muscular lesions.  相似文献   

14.
Abstract. The survival of Aeromonas salmonicida subsp. salmonicida was investigated in sea water under a variety of conditions. Survival in different types of microcosms (glass or dialysis bags); of bacteria grown under both in vivo and in vitro (broth culture) conditions; and in sterile and non-sterile sea water were compared. In all cases, survival was found to be of short duration (<10 days) and did not conflict with the previously stated obligate nature of the pathogen. The spread of furunculosis may depend less on its ability to survive in the environment than on its rate of shedding from infected fish and prevailing hydrographic conditions. Survival was extended and growth occurred in sterile sea water to which nutrients (tryptone soya broth) had been added. However, sea water obtained from beneath a commercial salmon cage, which would have been expected to be nutrient rich, did not prolong the survival of the pathogen. In vivo infectivity studies provided no evidence for the existence of unculturable but infective forms of A. salmonicida subsp. salmonicida which, therefore, validates the use of colony-forming units as a measure of survival.  相似文献   

15.
European sea bass (Dicentrarchus labrax) is a widely consumed marine fish in Mediterranean areas, and different farming techniques are applied for fish culturing to satisfy the growing demand for seafood. The aim of this study was to investigate the effects of conventional and organic feed on the quality of cultured European sea bass (Dicentrarchus labrax) collected during the growth period. The concentrations of ash, moisture, essential macro‐elements (Ca, K, Mg, Na and P), micro‐elements (Co, Cr, Cu, Fe, Mn, Ni and Zn) and toxic elements (As, Cd, and Pb) were determined in feeds and in fillets of cultured fish. The results were compared to those obtained from wild sea bass. Results showed that the differences between organic diet‐fed and conventional diet‐fed sea basses varied in relation to the specific element measured and the growth period. The former showed higher concentration of Fe, Mg and Cr, and lower Na content. The amount of P, Na, Fe, Cu and Cr in wild sea bass was significantly (P ≤ 0.05) higher than that found in farmed fish. The levels of toxic elements in cultured sea bass were always within the allowed limit for fishery products. Wild samples had significantly (P ≤ 0.05) higher content of arsenic and lead than farmed sea bass. Cultured fish represent a valuable dietary source of essential macro‐ and micro‐elements. Controlled rearing systems and feedings were related to a decrease in the presence of some toxic metals in cultured fish compared with wild fish.  相似文献   

16.
The location and cell damage caused by Vibrio anguillarum, the causative agent of classical vibriosis, within the developing gut of the newly hatched sea bass, Dicentrarchus labrax (L.), is unknown. A gnotobiotic sea bass model was used to investigate the early interactions of V. anguillarum with sea bass larvae. In the present study, germ‐free sea bass larvae were orally exposed to a V. anguillarum HI‐610 pathogen labelled with the green fluorescent protein (GFP‐HI‐610) and sampled at regular intervals. Pathogenic colonization of gut enterocytes was observed 2 h post‐exposure (p.e.) and onwards, whereas bacteria within the swim bladder were visualized 48 h p.e and onwards. Ultrastructural findings demonstrated direct bacterial contact with the host cell in the oesophageal mucosa and putative attachment to microvilli of mid‐ and hindgut enterocytes. The present findings form a starting point for studies assessing the impact of potential candidates (probiotics, prebiotics, antimicrobial peptides) to mitigate bacterial virulence.  相似文献   

17.
Yellowfin sea bream (Acanthopagrus latus) is an important economic fish, which is seriously threatened by various fish viruses. In this study, a cell line designated as ALL derived from the liver of yellowfin sea bream was developed and characterized. The cell line grew well in Dulbecco's modified Eagle's medium containing 10%–20% foetal bovine serum at 28°C. Amplification of the cytochrome B gene indicated that ALL cells originated from yellowfin sea bream. The modal chromosome number of ALL cells was 48. ALL cells were efficiently transfected with pEGFP-N3 plasmids, indicating the potential application of ALL cells in exogenous gene manipulation studies. ALL cells were susceptive to three main fish viruses, including viral haemorrhagic septicaemia virus (VHSV), red-spotted grouper nervous necrosis virus (RGNNV) and largemouth bass virus (LMBV). The replication of VHSV, RGNNV and LMBV in ALL cells was confirmed by quantitative real-time polymerase chain reaction, virus titre and transmission electron microscopy assays. Moreover, ALL cells could respond to VHSV, RGNNV and LMBV infections, as indicated by the differential expression of antiviral genes involving in the innate immune response. In conclusion, the newly established ALL cell line will be an excellent in vitro platform for the study of the virus–yellowfin sea bream interaction.  相似文献   

18.
Streptococcal infection ( Streptococcus iniae ) was diagnosed sequentially in two tanks of hybrid striped bass Morone saxatilis male × M. chrysops female (Sunshine bass) grown in a commercial freshwater recirculation facility in western Massachusetts. The pathogen was isolated in the laboratory, biochemically and morphologically characterized, and antibacterial sensitivities determined. Streptococcus iniae -induced lesions in the Sunshine bass were character at the gross and histopathological levels. Initial treatment with oxytetracycline was unsuccessful. Based on sensitivity results, enrofloxacin-medicated feed was dosed at 10 mg/kg body weight for 10 d, while a subsequent trial was conducted at 5 mg/kg body weight for 10 d. Mortality of fish subsided promptly following initiation of enrofloxacin therapy, yielding a final mortality in the initial tank of 10.83% (control tank 55.5%) and in the second tank of 16.97% (control tank 39.8%). Tissue enrofloxacin residues, detected via a microbiologic bioassay, revealed greater quantities and longer duration of residues in various tissues from the 5-mg as compared to the 10-mg trial, potentially the result of adverse feed palatability. Enrofloxacin appears to have excellent potential as an antibacterial agent for treating susceptible bacterial diseases of Sunshine bass.  相似文献   

19.
The effect of iron limitation, using the iron-chelating agent 2,2 dipyridyl, on the electrophoretic profiles of outer membrane proteins (OMPs) and extracellular products (ECPs) from 21 Pasteurella piscicida strains isolated from Europe and Japan was investigated. In addition, the effect of iron-limited and iron-surplus growth conditions on caseinase activity in culture supernatants of the pathogen was examined. The majority of P. piscicida strains, from Greece, Italy and France, cultured under iron-limited conditions, produced four novel OMPs (63 and three at and above 200 kDa). In contrast, iron-regulated outer membrane proteins were not induced in Japanese strains. Electrophoretic analysis of the ECPs from the pathogen grown under iron surplus and iron limitation revealed a large range of products and additional high molecular mass (MM) bands were evident under iron-limited conditions. When culture supernatants were analysed for their activity, most of the bacteria tested showed elevated activities under iron limited conditions. Finally, neither hydroxamate nor phenolate type siderophores could be detected with any of the chemical assays used.  相似文献   

20.
The objective of this study was to evaluate the effect of harvest practices and slaughter method on stress, quality and shelf life of whole fish (gilthead sea bream, European sea bass and red sea bream) towards the development of a humane slaughter practice. The use of hook and line resulted in significantly lower plasma cortisol and glucose in European sea bass and gilthead sea bream. Water temperature at harvest affected significantly the concentrations of stress indicators (plasma cortisol and glucose), mainly in European sea bass and gilthead sea bream. No significant differences were observed between the harvesting methods, regarding fish appearance, microbial spoilage and sensory scoring and subsequently shelf life. However, significantly lower hardness was recorded in fish harvested using the electric stunner, compared to the fish killed with slurry ice of the same harvesting period. Shelf life ranged between 13–18 days for gilthead seabream, 10–16 days for European sea bass and 10–14 days for red sea bream, depending on harvesting period.  相似文献   

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