Evaluation of candidate probiotic strains for gilthead sea bream larvae (Sparus aurata) using an in vivo approach

AIMS: The aim of this study was to evaluate the effect of six bacterial strains on gilthead sea bream larvae (Sparus aurata). METHODS AND RESULTS: Six bacterial strains isolated from well-performing live food cultures were identified by sequencing fragments of their 16s rDNA genome to the genus level as Cytophaga sp., Roseobacter sp., Ruergeria sp., Paracoccus sp., Aeromonas sp. and Shewanella sp. Survival rates of gilthead sea bream larvae transferred to seawater added these bacterial strains at concentrations of 6 +/- 0.3 x 10(5) bacteria ml(-1) were similar to those of larvae transferred to sterilized seawater and showed an average of 86% at 9 days after hatching, whereas, survival rates of larvae transferred to filtered seawater were lower (P < 0.05), and showed an average of 39%, 9 days after hatching. CONCLUSION: Several bacterial strains isolated from well-performing live food cultures showed a positive effect for sea bream larvae when compared with filtered seawater. SIGNIFICANCE AND IMPACT OF THE STUDY: The approach used in this study could be applied as an in vivo evaluation method of candidate probiotic strains used in the rearing of marine fish larvae.     

Harvesting <Emphasis Type="Italic">Chlorella minutissima</Emphasis> using cell coagulants

Twelve salts were tested for their ability to coagulate microalgae cells in cultures of Chlorella minutissima. The final aim was to develop an easy and efficient approach for harvesting microalgae biomass in dense cultures. Aluminum, ferric, and zinc salts coagulated C. minutissima cultures, while optimum concentration was 0.75 and 0.5 g L⁻¹ for sulfate and chloride salts, respectively. Aluminum salts were most efficient, but caused some cell lysis, which may render this approach inappropriate in some cases. Ferric and zinc salts were ranked second and third, respectively, according to their culture cell-coagulation efficiency. Ferric salts caused a change in the color of the cells, mainly at concentrations higher than 1 g L⁻¹. Zinc salts were less harmful for the microalgal cells, but an additional problem was observed with cell aggregates adhering to the walls of the glass test tubes. Selection of the appropriate coagulant is related to the purpose of the coagulation process.     

Environmental Conditioning of Skeletal Anomalies Typology and Frequency in Gilthead Seabream (Sparus aurata L., 1758) Juveniles

In this paper, 981 reared juveniles of gilthead seabream (Sparus aurata) were analysed, 721 of which were from a commercial hatchery located in Northern Italy (Venice, Italy) and 260 from the Hellenic Center for Marine Research (Crete, Greece). These individuals were from 4 different egg batches, for a total of 10 different lots. Each egg batch was split into two lots after hatching, and reared with two different methodologies: intensive and semi-intensive. All fish were subjected to processing for skeletal anomaly and meristic count analysis. The aims involved: (1) quantitatively and qualitatively analyzing whether differences in skeletal elements arise between siblings and, if so, what they are; (2) investigating if any skeletal bone tissue/ossification is specifically affected by changing environmental rearing conditions; and (3) contributing to the identification of the best practices for gilthead seabream larval rearing in order to lower the deformity rates, without selections. The results obtained in this study highlighted that: i) in all the semi-intensive lots, the bones having intramembranous ossification showed a consistently lower incidence of anomalies; ii) the same clear pattern was not observed in the skeletal elements whose ossification process requires a cartilaginous precursor. It is thus possible to ameliorate the morphological quality (by reducing the incidence of severe skeletal anomalies and the variability in meristic counts of dermal bones) of reared seabream juveniles by lowering the stocking densities (maximum 16 larvae/L) and increasing the volume of the hatchery rearing tanks (minimum 40 m³). Feeding larvae with a wide variety of live (wild) preys seems further to improve juvenile skeletal quality. Additionally, analysis of the morphological quality of juveniles reared under two different semi-intensive conditions, Mesocosm and Large Volumes, highlighted a somewhat greater capacity of Large Volumes to significantly augment the gap with siblings reared in intensive (conventional) modality.     

Uptake and processing of a Vibrio anguillarum bacterin in Artemia franciscana measured by ELISA and immunohistochemistry

Nauplii of Artemia franciscana were incubated in two different concentrations (undiluted and 1:9 in autoclaved sea water) of a divalent bacterin composed of two different serovars of Vibrio anguillarum. In order to investigate uptake and further processing of a bacterin in the live feed organism A. franciscana, immunohistochemistry was applied, visualising the presence of whole bacterial cells and antigens from the bacterin in individual nauplii. By using ELISA, it was shown that approximately 1.5-2-5 x 10(5) cells were incorporated into each Artemia under the conditions used. Maximum incorporation of cells was measured after 30 min, whereas after 60 min there was a decline to levels of 0.9-1.6 x 10(5) cells per Artemia. Immediately after incubation in the bacterin solution, the nauplii were transferred to a culture of the alga Isochrysis galbana, in order to simulate transfer of the nauplii to rearing tanks for fish larvae. From the ELISA, it could be concluded that the incorporated bacterial cells were excreted from the Artemia nauplii rapidly, however a large variation among different nauplii could be visualised by immunohistochemistry.     

Skin darkness is related to cortisol, but not MSH, content in post‐larval Solea senegalensis

In this study it was shown that dark coloured post‐larvae of Senegalese sole Solea senegalensis , at two different ages, had elevated cortisol concentrations compared with lighter coloured individuals. As melanophore‐stimulating hormone levels were not elevated in dark coloured fish, it is possible that this hormone may not be the main melanotropic hormone involved in stress‐related skin darkening.     

Food size selectivity of Artemia franciscana at three developmental stages

Food size selectivity was examined in Artemia franciscana metanaupliiat three different developmental stages. Clearance rates weredetermined in short-term experiments either by measuring thedecrease in concentration of live particles and plastic beads,or by measuring the radioactivity accumulated in animals thatgrazed ¹⁴C-labelled live particles. The maximum clearance rateofA.franciscana metanauplii increased during development andwas measured at 50–63 µl ind.^–1 h^–1,254 µl ind.^–1 h^–1 and 1.48–2.10 ml ind.^–1h^–1 in 2-, 4- and 7-day-old metanauplii, respectively.A preference for particles with a diameter of 4–8 µmwas observed at all three developmental stages. The abilityof A.franciscana metanauplii to graze bacterial particles wasalso demonstrated, although the efficiency in grazing such smallparticles was low compared to microalgae (28, 20 and 9% of themaximum clearance rate in 2-, 4- and 7-day-old metanauplii,respectively). Electron microscopy showed that the inter-setulardistance in antennae and thoracopods was 0.20 ± 0.07,0.16 ± 0.05 and 0.18 ± 0.04 µm in 2-, 4-and 7-day-old metanauplii, respectively, and accordingly independentof stage.     

Bioenergetic changes in the microalgal photosynthetic apparatus by extremely high CO2 concentrations induce an intense biomass production

Unicellular green alga Chlorella minutissima , grown under extreme carbon dioxide concentrations (0.036–100%), natural temperature and light intensities (Mediterranean conditions), strongly increase the microalgal biomass through photochemical and non-photochemical changes in the photosynthetic apparatus. Especially, CO₂ concentrations up to 10% enhance the density of active reaction centers (RC/CS_o), decrease the antenna size per active reaction center (ABS/RC), decrease the dissipation energy (DI_o/RC) and enhance the quantum yield of primary photochemistry (F_v/F_m). Higher CO₂ concentrations (20–25%) combine the above-mentioned photochemical changes with enhanced non-photochemical quenching of surplus energy, which leads to an enhanced steady-state fraction of 'open' (oxidized) PSII reaction centers (q_p), and minimize the excitation pressure of PSII (1 − q_p) under very high light intensities (approximately 1700 μmol m⁻² s⁻¹ maximal value), avoiding the photoinhibition and leading to an enormous biomass production (approximately 2500%). In conclusion, these extreme CO₂ concentrations – about 1000 times higher than the ambient one – can be easily metabolized from the unicellular green alga to biomass and can be used, on a local scale at least, for the future development of microalgal photobioreactors for the mitigation of the factory-produced carbon dioxide.