Isolation and characterization of bacteria associated with a syndrome disease of sea urchin Strongylocentrotus intermedius in North China

Sea urchin, Strongylocentrotus intermedius, transplanted from Japan in 1989, has been widely cultured along the coasts of Liaoning and Shandong Provinces and has become the dominant and most economically important maricultured species in North China. However, a lesion syndrome symptom of S. intermedius broke out frequently these years, showing lethargy in activities, blackish peristomial membrane and body well lesions, and brought about high mortality eventually. Six representative prominent bacterial strains were isolated from diseased sea urchin from September 2009 to January 2010. By means of API 20NE and 16S rRNA gene sequences analysis, isolates were identified as Shewanella aquimarina, Pseudoalteromonas tetraodonis, Vibrio shilonii, V. harveyi, V. fortis and V. splendidus. Bacterial challenge tests showed that their representative isolates were virulent to S. intermedius with LD₅₀ values ranging from 9.2 × 10⁴ to 3.4 × 10⁶ CFU/g body weight, among which S. aquimarina , V. fortis and P. tetraodonis were highly virulent, and the other three isolates showed moderate virulence. The results indicated that a variety of bacteria including Shewanella, Pseudoalteromonas and Vibrio were involved in the mortality of S. intermedius, and the six isolates were opportunistic pathogens of sea urchins. All isolates reported herein were sensitive to ampicillin, enrofloxacin, ofloxacin, doxycycline and florfenicol.     

Effects of diet and temperature on post‐settlement growth and survival of the short‐spined sea urchin Strongylocentrotus intermedius

Effects of diet and temperature on post‐settlement growth and survival of a sea urchin Strongylocentrotus intermedius, which fed on one of six mono‐cultured benthic diatoms, gametophytes of a brown alga Undaria pinnatifida, a green alga Ulvella lens plus mixed benthic diatoms, or no food at 6, 9, 12 and 15°C, were examined. Digestion efficiency (DE), ingestion rate (IR), excretion rate (ER) and digested cell content index (DCCI) of each diatom species were measured. Growth rates largely differed among the dietary treatments from 7 days post‐settlement. Sea urchins fed on a diatom Cocconeis sublittoralis showed considerably higher growth (7.85‐34.67 μm day^?1) than those fed on the other diets and high survival rates (100%). DCCI was also highest in C. sublittoralis. These results suggest that the quantity of diatom cell contents digested is an important factor affecting growth and survival of the sea urchins. Growth rates of sea urchins were higher in higher water temperatures, and significant differences in growth rate were observed between 6‐9 and 12‐15°C. ERs of C. sublittoralis were generally higher at higher temperatures, but DEs did not differ among the temperature treatments. Variations in growth rate among different water temperature appear to be caused by different food intakes of the urchins.     

Influence of stocking density and temperature on early development and survival of the purple sea urchin,Strongylocentrotus purpuratus (Stimpson, 1857)

We evaluated the effect of four densities (940, 1880, 3760, 7520 eggs cm^?2 and 0.5, 1, 2, 4 ind mL^?1 of embryos and larvae, respectively) and four temperatures (8, 11, 14, 17°C) on early growth and survival of the sea urchin Strongylocentrotus purpuratus. Prism‐stage length was significantly greater in embryos initially held at 940 and 1880 eggs cm^?2 than in those held at 3760 and 7520 eggs cm^?2. Larvae grew significantly faster and had significantly greater survival when reared at 0.5 or 1 ind mL^?1 than when held at 2 or 4 ind mL^?1. Embryos had greater survival at 11 and 14°C than at 8 and 17°C, whereas embryo length was significantly smaller at 8°C than at 11, 14 or 17°C. Larvae grew significantly slower at 8°C than at 11, 14 or 17°C, whereas survival was significantly reduced at 8 and 17°C compared with 11 and 14°C. Per cent survival from prism to metamorphic competency in the best treatments was 48.9 ± 2.2% and 50.0 ± 3.6% (mean ± SE) for the 1 ind mL^?1 and 11°C treatments, respectively. On the basis of these results, for rearing of S. purpuratus under static conditions, we recommend that fertilized eggs and larvae be held at ≤1880 eggs cm^?2 and ≤1 ind mL^?1, respectively, and at 11–14°C.     

Determination of a novel bitter amino acid, pulcherrimine, in the gonad of the green sea urchin Hemicentrotus pulcherrimus

SUMMARY: A novel bitter amino acid, pulcherrimine was analyzed by reverse-phase high-performance liquid chromatography using a linear gradient elution consisting of methanol/aqueous/acetic acid and methanol/acetic acid mixture, after being labeled with dimethylaminoazobenzenesulfonyl chloride (Dabs-Cl). Linearity of the calibration curve for pulcherrimine was high ( r = 0.994) in the tested range from 0 to 4 μg/mL. By this method, the pulcherrimine content in the ovary and testis of the green sea urchin Hemicentrotus pulcherrimus , collected in the sea off Iwaki in November 1998, was analyzed. The mean pulcherrimine content in the ovary was 1.37 mg/100 g. In contrast, no pulcherrimine was detected in all testes examined. Correlation between the bitterness and the pulcherrimine content was statistically significant. These results strongly support that pulcherrimine is a bitter principle in the green sea urchin ovaries.     

The effect of anti-Paramoeba antibodies on Paramoeba sp., the causative agent of amoebic gill disease

The effect of anti- Paramoeba antibodies produced in sheep on Paramoeba sp., the causative agent of amoebic gill disease (AGD), was tested. Incubation of Paramoeba sp. with antiserum at a concentration of 20% or more effectively coated the Paramoeba with antibodies. The antiserum had no effect on Paramoeba sp. survival in vitro or on its ability to cause AGD in Atlantic salmon.     

Feeding time,frequency and ration affect growth and energy allocation in young adults of the sea urchin Lytechinus variegatus

In a 62‐day growth trial, wild caught Lytechinus variegatus (ca. 18 g, 35 ± 2 mm dia.) were fed as follows: (i) ration equivalent to 3% of average body weight proffered once daily in the morning (AM treatment); (ii) ration equivalent to 3% of average body weight proffered once daily in the evening (PM treatment); (iii) ration equivalent to 1.5% of average body weight proffered twice daily (every 12 h, AM/PM treatment); (iv) ration equivalent to 3% of average body weight proffered in the evening on alternate days (EODSR/PM); or (v) ration equivalent to 6% of average body weight proffered in the evening on alternate days (EODDR/PM). At 30 and 62 days, wet weight gain among urchins fed daily was significantly higher than that of urchins fed on alternate days, regardless of ration size. At 62 days, reduced gonad dry matter production and gonad index were observed among individuals fed on alternate days, regardless of ration size. At 30 and 62 days, feed conversion ratio was optimized among individuals fed daily and in the EODSR/PM treatment. These data suggest that, for culture of L. variegatus, daily feeding will optimize growth, gonad production and feed efficiency regardless of ration size or time of feeding.     

Production of salmonid amoebic gill disease by exposure to Paramoeba sp. harvested from the gills of infected fish

Atlantic salmon, S almo salar L., were exposed to different concentrations of Paramoeba sp. The lowest concentration which induced amoebic gill disease (AGD) was 230 Paramoeba sp. L^–1 and the severity of infection increased with increasing concentration. The concentration of Paramoeba sp. positively correlated with the number of gill lesions ( R ² > 0.7). This study provides evidence that Paramoeba sp. is the causative agent of AGD and describes an experimental model that enables the severity of the induced disease to be controlled.     

Observations of Dermocystidium sp. infections in bullheads, Cottus gobio L., from a river in southern England

Bullheads, Cottus gobio, with macroscopic external cysts on the skin and fins measuring up to 3 mm in diameter were detected in the River Allen and its tributaries in southern England between 1992 and 1998. The prevalence of these cysts was up to 50% at some sites. Examination of cyst contents revealed the presence of numerous spores, typical of the genus Dermocystidium, measuring 8 microm in diameter. The parasite developed within well-defined cysts, which were located in the hypodermal connective tissues of the host. No cysts were present on the fins of any of the fish examined. Histological examination revealed a cyst wall consisting of an inner layer of dense eosinophilic material similar to that reported for Dermocystidium spp. forming coenocytic hyphae. No evidence was found of systemic infection or hyphal formation. Spores contained a prominent refractile body, which gave a weakly positive reaction for polysaccharides with the periodic-acid Schiff reaction and was positively stained with acidic dyes. Several examples of ruptured cysts were seen in histological sections and in some of these cases the host epithelial layer was breached, allowing release of the spores to the environment. Morphological features of, and host response towards, the Dermocystidium sp. in bullheads are compared with similar infections in salmonids and other freshwater fish species.     

Neoparamoeba branchiphila n. sp., and related species of the genus Neoparamoeba Page, 1987: morphological and molecular characterization of selected strains

A total of 18 Neoparamoeba strains were characterized both morphologically and using the SSU rRNA gene sequences as molecular markers. Nine were isolated from gills of farmed Atlantic salmon, Salmo salar L., six from sediments sampled in areas of sea-cage farms and three from net material of sea-cages. The newly obtained sequences extended substantially the dataset of Neoparamoeba strains available for phylogenetic analyses, which were used to infer taxonomic relatedness among 32 strains morphologically assigned to this genus. In addition to the N. pemaquidensis and N. aestuarina clades, phylogenetic analyses clearly distinguished a third clade with sequences from six strains. Members of this clade are characterized as representatives of a new species, N. branchiphila n. sp. The diagnostic primers for the identification of this species are introduced.     

Changes in the innate immune response of Atlantic salmon, Salmo salar L., exposed to experimental infection with Neoparamoeba sp

An experiment was conducted to determine the effect of Neoparamoeba sp. infection on the innate immune responses of Atlantic salmon. Atlantic salmon were experimentally infected with Neoparamoeba sp. and serially sampled 0, 1, 4, 6, 8 and 11 days post-exposure (dpe). Histological analysis of infected fish gill arches identified the presence of characteristic amoebic gill disease lesions as early as 1 dpe with a steady increase in the number of affected gill filaments over time. Immune parameters investigated were anterior kidney phagocyte function (respiratory burst, chemotaxis and phagocytosis) and total plasma protein and lysozyme. In comparison with non-exposed control fish basal respiratory burst responses were suppressed at 8 and 11 dpe, while phorbol myristate acetate-stimulated activity was significantly suppressed at 11 dpe. Variable differences in phagocytic activity and phagocytic rate following infection were identified. There was an increase in the chemotactic response of anterior kidney macrophages isolated from exposed fish relative to control fish at 8 dpe. Total protein and lysozyme levels were not affected by Neoparamoeba sp. exposure.     

Effects of dietary fish oil and soya bean lecithin on gonad index,colour and biochemical composition of the purple sea urchin,Strongylocentrotus purpuratus (Stimpson 1857)

The effects of increasing soya bean lecithin (SL) levels (0%, 2% and 4% of diet dry matter, DM) at two fish oil (FO) levels (0% and 3% DM) on gonad index, colour and biochemical composition of Strongylocentrotus purpuratus were evaluated using a 3 × 2 factorial design with six iso‐nitrogenous formulated diets. All diets generated an increase in gonad index of more than 9%. Dietary FO had a positive effect on diet digestibility and accumulation of proteins in the gonad. Soy lecithin did not affect gonad composition, but improved its coloration, decreased gonad feed conversion ratio and increased gonad protein efficiency. There was a clear effect of diets on sea urchins gonad protein, ash and fatty acid content, in particular, decreasing the n‐3/n‐6 ratio associated with a decrease in 20:5n‐3 when SL levels increased. Nevertheless, 22:6n‐3 levels increased by week 12 in all treatments, and diets with SL increased the content of 20:4n‐6 in gonads. A decrease in bitter‐tasting amino acids such as histidine, cysteine, valine and methionine was observed in all treatments with a concomitant increase in isoleucine levels. We recommend using a diet containing 3% of FO and 2% of SL to increase food consumption, diet digestibility, improving marketable gonad colour and an increasing gonad n‐6 fatty acids and C22:6n‐3. In addition, the present study paves the way for future research in the use of FO and SL as an additive in diets for S. purpuratus towards the goal of increasing gonad size and nutritional quality.     

Evaluation of dried macrophytes as an alternative diet for the rearing of the sea urchin Paracentrotus lividus (Lamarck, 1816)

The aim of this work was to assess the potential use of different dried macroalgae as food in the rearing of Paracentrotus lividus. Growth, consumption and food conversions were compared in adult sea urchins fed with fresh or dried thalli of four macroalgae species. Six experimental diets were tested: (a) fresh Palmaria palmata; (b) fresh Saccharina latissima; (c) dry P. palmata; (d) dry S. latissima; (e) dry Laminaria digitata and (f) dry Grateloupia turuturu. Linear growth rates were similar for all treatments. Specific growth rate was higher in sea urchins fed with fresh P. palmata, but no difference was found between animals fed with fresh S. latissima and those fed with dried diets. Regarding daily food consumption (DFC), sea urchins consumed the same amount of dried macroalgae as fresh but exhibited a higher food conversion efficiency (FCE) when fed with fresh P. palmata. However, this FCE was only significantly higher when compared to sea urchins fed with dry L. digitata. Dried G. turuturu is not a suitable diet due to its rapid degradation after rehydration. The results suggest that P. lividus adults can be reared on dried macroalgae thalli without detriment to their somatic growth, especially over short periods. The low cost of feeding sea urchins with this diet could help small shellfish farmers to diversify their production into echinoculture.     

Isolation of lymphocystis disease virus from sole, Solea senegalensis Kaup, and blackspot sea bream, Pagellus bogaraveo (Brunnich)

Two viruses were isolated from cultured sole, Solea senegalensis, and wild blackspot sea bream, Pagellus bogaraveo, and preliminarily characterized as lymphocystis disease viruses (LCDVs). Viral isolates were characterized by morphological, biochemical and biophysical properties. In addition, the susceptibility of four fish cell lines was also tested. LCDV isolates developed cytopathic effects on the SAF-1 cell line at 5 and 6 days post-infection and reached titres of 10(6) TCID50 mL(-1). The antigenic and structural protein analysis of the two new LCDV isolates showed identical profiles to that obtained for LCDV strain Leetown NFH (ATCC VR-342), used as a reference viral strain, and for an LCDV isolate collected from gilt-head sea bream, Sparus aurata, cultured in southern Spain. Molecular confirmation was performed by polymerase chain reaction. Specific primers for LCDV produced a 270-bp DNA fragment, the expected size for LCDV.     

The performance of sea trout,Salmo trutta L., stocks from the Burrishoole system western Ireland, 1970–1994.

Abstract Since 1975, with partial recoveries in 1979 and 1984–1986, the numbers of finnock and adult sea trout, Salmo trutta L., in the Burrishoole system, western Ireland, have declined. The population decreased sharply from 1987, and collapsed in 1989 and 1990. Finnock were virtually absent in 1989 and one-year maidens equally so in 1990. Sea trout smolt recruitments from 1992 to 1994 were the lowest recorded since 1970. The modal fork length of finnock decreased from 28 cm in 1987 to 26 cm in 1990. Up until 1989, the percentage return of smolts, as finnock, ranged from 11.4 to 32.4%. In 1989 a minimum was reached at 1.5%, with values of 10.0, 3.7, 7.4 and 9.9% from 1990 to 1994. Marine survival of smolts, to total first return to fresh water, historically ranged from 19 to 66%. Survival dropped to 1.8% in 1989. Observed changes in the structure of the sea trout population and the collapse in marine survival suggest that the current west of Ireland sea trout problem is based in the marine habitat.     

14株患病水生动物气单胞菌属细菌的分离鉴定与最佳治疗药物筛选试验

从患病甲鱼、河蟹、牛蛙、鲫体内共分离到14株气单胞菌属细菌,对它们进行了生化鉴定和药敏试验。根据药敏试验结果,选取1株嗜水气单胞菌(J50503-1)和1株温和气单胞菌(NT712-10)作了最小抑菌浓度试验和人工感染及治疗试验。结果表明,氟苯尼考对上述细菌有很好的防治作用。