Settling abalone veliger larvae in a free-swimming microalgal culture

In the current abalone hatchery in China, insufficient diatoms on vertically placed corrugated pvc plates at later stage often could not support the growth of postlarvae up to the stage that they can feed on live macroalgae. As a result, stripping the spats (35 mm) off by anaesthetization and switching the diet from live diatoms to artificial powdered diet in combination has to be performed in most of the abalone farms. This manipulation normally leads to more than 50% mortality. Here we report the direct use of the unicellular green alga Platymonas helgolandica Kylin var. tsingtaoensis as a potential alga to be used to settle the veliger larvae of the Pacific abalone Haliotis discus hannai and to feed the postlarvae. Settlement rate of 2-day-old veliger larvae in mono culture of P helgolandica could be as high as 92% ( +/- 4.2%) on day 10 in small scale trials, higher than that in the selected benthic diatom strain (53.6% +/- 12.7%) when settled in the water in which bacteria propagation was controlled by treatment of 2 ppm of benzylpenicillinum calcium and streptomycin sulfate. Postlarvae fed solely on P. helgolandica or the selected benthic diatom Navicula-2005-A grew at rates of 40.1 ( +/- 1.9) and 45.8 (+/- 13.4) mu m day(-1), respectively, when raised at 22 degrees C until day 50 postfertilization. P. helgolandica was shown to have distinct diurnal settling rhythm characterized with a peak of settled cells in the middle of the night for cell division and a peak of free-swimming cells in the middle of the day. High density of attached P. helgolandica cells on the inner surface of the culture facility in the night fits the nocturnal feeding behavior of the abalone spats. Judged by the promising larvae settling rate, growth and survival rates of the postlarvae fed with this alga, the free-swimming micro-green alga P. helgolandica constitutes a potential species for settling the veliger larvae and for supporting the growth of postlarvae as well. (c) 2006 Published by Elsevier B.V.

Growth, survival and immune activity of scallops, Chlamys farreri Jones et Preston, compared between suspended and bottom culture in Haizhou Bay, China

We examined the growth, survival and immune response of the scallop, Chlamys farreri, during a 1-year period in deep water of Haizhou Bay. Scallops were cultured using two methods: (1) in lantern nets at a 5 m depth and (2) in a bottom culture system (sleeves) on the seabed at about a 25 m depth. Shell heights, meat dry weight and immune activities in the haemolymph (superoxide dismutase and myeloperoxidase) were measured bimonthly or quarterly from July 2007 to June 2008. Survival was measured at the end of the study and environmental parameters in the experimental layers were monitored during the experiment. The growth and immune activities of scallops were lower when the water temperature was high, which was consistent with the main mortality occurring in summer. The growth and immunity of scallops were higher in the suspended culture than in the bottom culture during the experiment, with the exception of shell growth during the last study period. Survival of scallops in the suspended culture (54.6 +/- 12.3%) was significantly lower than that in the bottom culture (86.8 +/- 3.5%) at the end of this study. We conclude from our results that the high mortality of C. farreri can be prevented by culturing them in a bottom culture system before November of the first year, and then transferring them to a suspended culture to improve scallop production.

Bioremediation potential of the macroalga Gracilaria lemaneiformis (Rhodophyta) integrated into fed fish culture in coastal waters of north China

Fed fish farms produce large amounts of wastes, including dissolved inorganic nitrogen and phosphorus. In China, fish mariculture in coastal waters has been increasing since the last decade. However, there is no macroalgae commercially cultivated in north China in warm seasons. To exploit fish-farm nutrients as a resource input, and at the same time to reduce the risk of eutrophication, the high-temperature adapted red alga Gracilaria lemaneiformis (Bory) Dawson from south China was co-cultured with the fish Sebastodes fuscescens in north China in warm seasons. Growth and nutrient removal from fish culture water were investigated in laboratory conditions in order to evaluate the nutrient bioremediation capability of G. lemaneiformis. Feasibility of integrating the seaweed cultivation with the fed fish-cage aquaculture in coastal waters of north China was also investigated in field conditions. Laboratory seaweed/fish co-culture experiments showed that the seaweed was an efficient nutrient pump and could remove most nutrients from the system. Field cultivation trials showed that G. lemaneiformis grew very well in fish farming areas, at maximum growth rate of 11.03% day(-1). Mean C, N, and P contents in dry thalli cultured in Jiaozhou Bay were 28.9 +/- 1.1%, 4.17 +/- 0.11 % and 0.33 +/- 0.01 %, respectively. Mean N and P uptake rates of the thalli were estimated at 10.64 and 0.38 mu mol g(-1) dry weight h(-1), respectively. An extrapolation of the results showed that a 1-ha cultivation of the seaweed in coastal fish fanning waters would give an annual harvest of more than 70 t of fresh G. lemaneiformis, or 9 t dry materials; 2.5 t C would be produced, and simultaneously 0.22 t N and 0.03t P would be sequestered from the seawater by the seaweed. Results indicated that the seaweed is suitable as a good candidate for seaweed/fish integrated mariculture for bioremediation and economic diversification. The integration can benefit economy and environment in a sustainable manner in warm seasons in coastal waters of north China. (c) 2005 Elsevier B.V. All rights reserved.

Isolation and characterization of a marine magnetotactic spirillum axenic culture QH-2 from an intertidal zone of the China Sea

Magnetotactic bacteria (MTB) are ubiquitous in aquatic habitats. Because of their fastidious requirements for growth conditions, only very few axenic MTB cultures have been obtained worldwide. In this study, we report a novel marine magnetotactic spirillum axenic culture, designated as QH-2, isolated from the China Sea. It was able to grow in semi-solid or liquid chemically defined medium. The cells were amphitrichously flagellated and contained one single magnetosome chain with an average number of 16 magnetosomes per cell. Phosphate and lipid granules were also observed in the cells. Both rock magnetism and energy-dispersive X-ray spectroscopy characterizations indicated that the magnetosomes in QH-2 were single-domain magnetites (Fe3O4). QH-2 cells swam mostly in a straight line at a velocity of 20-50 mu m/s and occasionally changed to a helical motion. Unlike other magnetotactic spirilla. QH-2 cells responded to light illumination. As a consequence of illumination, the cells changed the direction in which they swam from parallel to the magnetic field to antiparallel. This response appears to be similar to the effect of an increase in [O-2]. Analysis of the QH-2 16S rRNA sequence showed that it had greater than 11% sequence divergence from freshwater magnetotactic spirilla. Thus, the marine QH-2 strain seems to be both phylogenetically and magnetotactically distinct from the freshwater Magnetospirillum spp. studied previously. (C) 2010 Elsevier Masson SAS. All rights reserved.

A new three-phase culture method for Manila clam, Ruditapes philippinarum, farming in northern China

Studies on reproduction, hatchery management, and culture of Manila clams Ruditapes philippinarum were carried out in an attempt to optimize their culture conditions and techniques. Results from these studies led to the development of a three-phase culture method for Manila clam farming in northern China. The key components of the new method were: 1) early spawning and over-wintering indoors (greenhouse); 2) optimized larval culture conditions and techniques; 3) juvenile rearing in shallow, fertilized nursery ponds; 4) optimized stocking size and density and substrate for mudflat grow out. Broodstock were maturated indoors for a month from early April to early May. Primarily because of higher water temperatures in the greenhouse the clams spawned more than one month earlier than in the natural environment. From May to July, juveniles were reared for 1-2 months indoors to a size of 2.0-3.0 mm in shell length before being moved to outdoor, pre-disinfected, nursery ponds. Juveniles were then reared in the nursery ponds for one month to about 1.0 cm before being transferred to the mudflat for grow out. Juvenile clams in nursery ponds grew considerably faster than in the natural environment probably because of higher temperatures and more abundant natural food. During grow out, the clams were reared for 4-7 months until they reached a market size (3.0-3.3 cm). Juveniles produced after August were over-wintered in the greenhouse in which the water temperature was about 3 degrees C higher than that of the outdoor environment. Juveniles grew at an average rate of > 20 mu m day(-1), while in the natural environment no growth was observed during winter because of low temperatures. Juveniles in the greenhouse grew to 2-3 mm by the following March before being moved into outdoor nursery ponds. The three-phase culture method not only shortened the production period from spawn to market size from 24-36 months to about 10-14 months, but also prolonged the spawning season from 2 to 7 months, resulting in increased production of seed and market-size clams. Compared with the traditional method, the new method could increase the yield of market-size clams by 10-11 times, and increase the profit per ha mudflat by as much as 124 times and the profit per kg market-size clams produced by 13 times. (c) 2006 Elsevier B.V. All rights reserved.

Analysis for Flavonoids in Bee Pollens by Capillary Electrophoresis

A capillary electrophoresis (CE) method has been developed for the determination of six bioactive flavonoids that are commonly found in health foods: hesperidin, hyperin, isorhamnetin, kaempferol, quercetin and rutin. The effects of several parameters, such as pH, buffer concentration, separation voltage and UV detector wavelength, were investigated to find the optimal conditions. Using a HbBCh-NaiB-iO? buffer (pH 9.2), the analytes can be separated within 8 min. The relative standard deviations of migration times in eight injections were between 0.77% and 0.93%, and those of the peak areas ranged from 3.8% to 8.6%. A high reproducibility and excellent linearity was observed over two orders of magnitude, with detection limits (S/N = 3) ranging from 0.34ug ml to 2.9ug ml for all the six analytes. Recoveries ranged from 80.4 % to 113.9 %. The new method is simple, reproducible and sensitive. No solid phase extraction for sample pretreatment is necessary. Analysis results are accurate in application to bee pollens.

Study of a new derivatizing reagent that improves the analysis of amino acids by HPLC with fluorescence detection: application to hydrolyzed rape bee pollen

A simple and sensitive method for evaluating the chemical compositions of protein amino acids, including cystine (Cys)(2) and tryptophane (Try) has been developed, based on the use of a sensitive labeling reagent 2-(11H-benzo[alpha]-carbazol-11-yl) ethyl chloroformate (BCEC-Cl) along with fluorescence detection. The chromophore of the 1,2-benzo-3,4-dihydrocarbazole-ethyl chloroformate (BCEOC-Cl) molecule was replaced with the 2-(11H-benzo[alpha]-carbazol-11-yl) ethyl functional group, yielding the sensitive fluorescence molecule BCEC-Cl. The new reagent BCEC-Cl could then be substituted for labeling reagents commonly used in amino acid derivatization. The BCEC-amino acid derivatives exhibited very high detection sensitivities, particularly in the cases of (Cys)(2) and Try, which cannot be determined using traditional labeling reagents such as 9-fluorenyl methylchloroformate (FMOC-Cl) and ortho-phthaldialdehyde (OPA). The fluorescence detection intensities for the BCEC derivatives were compared to those obtained when using FMOC-Cl and BCEOC-Cl as labeling reagents. The ratios I (BCEC)/I (BCEOC) = 1.17-3.57, I (BCEC)/I (FMOC) = 1.13-8.21, and UVBCEC/UVBCEOC = 1.67-4.90 (where I is the fluorescence intensity and UV is the ultraviolet absorbance). Derivative separation was optimized on a Hypersil BDS C-18 column. The detection limits calculated from 1.0 pmol injections, at a signal-to-noise ratio of 3, ranged from 7.2 fmol for Try to 8.4 fmol for (Cys)(2). Excellent linear responses were observed, with coefficients of > 0.9994. When coupled with high-performance liquid chromatography, the method established here allowed the development of a highly sensitive and specific method for the quantitative analysis of trace levels of amino acids including (Cys)(2) and Try from bee-collected pollen (bee pollen) samples.

Enhanced anthocyanin production by repeated-batch culture of strawberry cells with medium shift

Repeated-batch cultures of strawberry cells (Fragaria ananassa cv. Shikinari) subjected to four medium-shift procedures (constant LS medium, constant B5 medium, alternation between LS and B5 starting from LS and alternation between LS and B5 starting from B5) were investigated for the enhanced anthocyanin productivity. To determine the optimum period for repeated batch cultures, two medium-shift periods of 9 and 14 days were studied, which represent the end of the exponential growth phase and the stationary phase. By comparison with the corresponding batch cultures, higher anthocyanin productivity was achieved for all the repeated-batch cultures at a 9-day medium-shift period. The average anthocyanin productivity was enhanced 1.7-and 1.76-fold by repeated-batch cultures in constant LS and constant B5 medium at a 9-day shift period for 45 days, respectively. No further improvement was observed when the medium was alternated between LS (the growth medium) and B5 (the production medium). Anthocyanin production was unstable at a 14-day shift period regardless of the medium-shift procedures. The results show that it is feasible to improve anthocyanin production by a repeated-batch culture of strawberry cells.

Formulation of a basal medium for primary cell culture of the marine sponge Hymeniacidon perleve

Marine sponge cell culture is a potential route for the sustainable production of sponge-derived bioproducts. Development of a basal culture medium is a prerequisite for the attachment, spreading, and growth of sponge cells in vitro. With the limited knowledge available on nutrient requirements for sponge cells, a series of statistical experimental designs has been employed to screen and optimize the critical nutrient components including inorganic salts (ferric ion, zinc ion, silicate, and NaCl), amino acids (glycine, glutamine, and aspartic acid), sugars (glucose, sorbitol, and sodium pyruvate), vitamin C, and mammalian cell medium (DMEM and RPMI 1640) using MTT assay in 96-well plates. The marine sponge Hymeniacidon perleve was used as a model system. Plackett-Burman design was used for the initial screening, which identified the significant factors of ferric ion, NaCl, and vitamin C. These three factors were selected for further optimization by Uniform Design and Response Surface Methodology (RSM), respectively. A basal medium was finally established, which supported an over 100% increase in viability of sponge cells.