Nitrate and phosphate supplementation to increase toxin production by the marine dinoflagellate Alexandrium tamarense

Alexandrium tamarense toxins have great value in biotechnology research as well as important in connection with shellfish poisoning. The influence of nitrate or nitrate and phosphate supplementation on cell biomass and toxin content were investigated in batch cultures. When cultures at low nitrate (88.2 mu M NaNO3) Were supplemented with 793.8 mu M NaNO3 at day 10 the cell density and cellular toxin contents were increased by 6-29% and 20-76%, respectively, compared with controls, and maximal values were 43,600 cells/ml (day 38) and 0.91 pg/cell (day 31). Supplementation with nitrate at day 14 or with nitrate and phosphate at day 10/14 to the cultures did not increase the cell density compared with the non-supplemented middle nitrate or high phosphate (108 mu M NaH2PO4) cultures, respectively, but increased the cellular toxin contents by an average of 52%. The results showed that supplementation with nitrate or with nitrate and phosphate at different growth phases of the cultures increased toxin yield by an average of 46%. Supplementation with nitrate at selected times to maintain continuous low level of nitrate might contribute to the effective increase of toxin yield of A. tamarense. (c) 2005 Elsevier Ltd. All rights reserved.

Effects of solar UV radiation on morphology and photosynthesis of filamentous cyanobacterium Arthrospira platensis

To study the impact of solar UV radiation (UVR) (280 to 400 nm) on the filamentous cyanobacterium Arthrospira (Spirulina) platensis, we examined the morphological changes and photosynthetic performance using an indoor-grown strain (which had not been exposed to sunlight for decades) and an outdoor-grown strain (which had been grown under sunlight for decades) while they were cultured with three solar radiation treatments: PAB (photosynthetically active radiation [PAR] plus UVR; 280 to 700 nm), PA (PAR plus UV-A; 320 to 700 nm), and P (PAR only; 400 to 700 nm). Solar UVR broke the spiral filaments of A. platensis exposed to full solar radiation in short-term low-cell-density cultures. This breakage was observed after 2 h for the indoor strain but after 4 to 6 h for the outdoor strain. Filament breakage also occurred in the cultures exposed to PAR alone; however, the extent of breakage was less than that observed for filaments exposed to full solar radiation. The spiral filaments broke and compressed when high-cell-density cultures were exposed to full solar radiation during long-term experiments. When UV-B was screened off, the filaments initially broke, but they elongated and became loosely arranged later (i.e., there were fewer spirals per unit of filament length). When UVR was filtered out, the spiral structure hardly broke or became looser. Photosynthetic 0, evolution in the presence of UVR was significantly suppressed in the indoor strain compared to the outdoor strain. UVR-induced inhibition increased with exposure time, and it was significantly lower in the outdoor strain. The concentration of UV-absorbing compounds was low in both strains, and there was no significant change in the amount regardless of the radiation treatment, suggesting that these compounds were not effectively used as protection against solar UVR. Self-shading, on the other hand, produced by compression of the spirals over adaptive time scales, seems to play an important role in protecting this species against deleterious UVR. Our findings suggest that the increase in UV-B irradiance due to ozone depletion not only might affect photosynthesis but also might alter the morphological development of filamentous cyanobacteria during acclimation or over adaptive time scales.

Ecophysiological characteristics of four intertidal marine macroalgae during emersion along Shantou coast of China, with a special reference to the relationship of photosynthesis and CO2

Intertidal marine macroalgae experience periodical exposures during low tide due to their zonational distribution. The duration of such emersion leads to different exposures of the plants to light and aerial CO2, which then affect the physiology of them to different extents. The ecophysiological responses to light and CO2 were investigated during emersion in two red algae Gloiopeltis furcata and Gigartina intermedia, and two brown algae Petalonia fascia and Sargassum hemiphyllum, growing along the Shantou coast of China. The light-saturated net photosynthesis in G. furcata and P. fascia showed an increase followed by slightly desiccation, whereas that in G. intermedia and S. hemiphyllum exhibited a continuous decrease with water loss. In addition, the upper-zonated G. furcata and P. fascia, exhibited higher photosynthetic tolerance to desiccation and required higher light level to saturate their photosynthesis than the lower-zonated G. intemedia and S. hemiphyllum. Desiccation had less effect on dark respiration in these four algae compared with photosynthesis. The light-saturated net photosynthesis increased with increased CO2 concentrations, being saturated at CO2 concentrations higher than the present atmospheric level in G. furcata, G. intermedia and S. hemiphyllum during emersion. It was evident that the relative enhancement of photosynthesis by elevated CO, in those three algae increased, though the absolute values of photosynthetic enhancement owing to CO2 increase were reduced when the desiccation statuses became more severe. However, in the case of desiccated P. fascia (water loss being greater than 20 %), light saturated net photosynthesis was saturated with current ambient atmospheric CO2 level. It is proposed that increasing atmospheric CO2 will enhance the daily photosynthetic production in intertidal macroalgae by varied extents that were related to the species and zonation.

Photoregulated or energy dependent process of hormogonia differentiation in Nostoc sphaeroides Kutzing (Cyanobacterium)

Hormogonium, which was thought to play an important role in the dispersal and survival of these microorganisms in their natural habitats, is a distinguishable developmental stage of heterocystous cyanobacteria. The present study examined the effects of different light conditions and sugars on the differentiation of Nostoc sphaeroides Kutzing to the hormogonia stage. Results showed that differentiation of hormogonia was light dependent in the absence of sugar, but that close to 100% of cyanobacteria differentiated to hormogonia in the presence of glucose or sucrose, irrespective of the light conditions. This differentiation was inhibited, even in the presence of sugars, upon application of an inhibitor of respiration. Following the testing of different sugars, the effects of different lights were examined. It was found that 5 10 μ mol.m(-2)• s(-1) photon flux density was optimal for hormogonia differentiation. One hundred percent differentiation was obtained with white light irradiation, in contrast with irradiation with green light (80% differentiation) and red light (0-10% differentiation). Although they showed different efficiencies in inducing hormogonia differentiation in N. sphaeroides, the green and red radiation did not display antagonistic effects. When the additional aspect of time dependence was investigated through the application of different light radiations and an inhibitor of protein synthesis, it was found that the initial 6 h of the differentiation process was crucial for hormogonia differentiation. Taken together, these results show that hormogonia differentiation in N. sphaeroides is either a photoregulated or an energy dependent process.

Reactive oxygen species and antioxidant enzymes activity of Anabaena sp PCC 7120 (Cyanobacterium) under simulated microgravity

It was found that reactive oxygen species in Anabaena cells increased under simulated microgravity provided by clinostat. Activities of intracellular antioxidant enzymes, such as superoxide dismutase, catalase were higher than those in the controlled samples during the 7 days' experiment. However, the contents of gluathione, an intracellular antioxidant, decreased in comparison with the controlled samples. The results suggested that microgravity provided by clinostat might break the oxidative/antioxidative balance. It indicated a protective mechanism in algal cells, that the total antioxidant system activity increased, which might play an important role for algal cells to adapt the environmental stress of microgravity. (C) 2004 Elsevier Ltd. All rights reserved.

Characterization of diurnal photosynthetic rhythms in the marine diatom Skeletonema costatum grown in synchronous culture under ambient and elevated CO2

Photosynthetic performance was examined in Skeletonema costatum (Greville) Cleve. under 12: 12-h light: dark (LD) cycle at ambient CO2 (350 muL L-1) and elevated CO2 (1000 muL L-1). At ambient CO2, the cellular chlorophyll a content, the light-saturated photosynthetic rate (P-m), the initial slope of the light saturation curves ( a), the photochemical efficiency of PSII (F-v/F-m), the apparent carboxylating efficiency (ACE) and the photosynthetic affinity for CO2 [1/K-m (CO2)] all showed rhythmical changes with different amplitudes during the light period. The P-m had similar changing pattern in the light period with the ACE and 1/K-m (CO2) rather than with the alpha and F-v/F-m, indicating that rhythmical changes of photosynthetic capacity may be mainly controlled by the activity of C- reduction associated with CO2 uptake during the light period. The CO2 enrichment reduced the ACE and the affinity to CO2, and increased the a, cellular chlorophyll a content and P m based on cell number. By contrast, the changing patterns of all photosynthetic parameters examined here during the light period had almost the same for cells grown at ambient CO2 and elevated CO2, suggesting that the photosynthetic rhythms of S. costatum are not affected by CO2 enrichment.

Broad host range plasmid-based gene transfer system in the cyanobacterium Gloeobacter violaceus which lacks thylakoids

Gloeobacter violaceus, a cyanobacterium lack of thylakoids, is refractory to genetic manipulations because its cells are enveloped by a thick gelatinous sheath and in colonial form. In this study, a large number of single cells were obtained by repeated pumping with a syringe with the gelatinous sheath removed. And an exogenous broad host range plasmid pKT210 was conjugatively transferred into G. violaceus. Analyses with dot-blot hybridization and restriction mapping showed that the exogenous plasmid pKT210 had been introduced into G. violaceus and stably maintained with no alteration in its structure. pKT210 extracted from G. violaceus exconjugants could be transformed into the mcr - mrr - E. coli strain DH10B but not the mcr(+) mrr(+) strain DH5alpha, which suggests that a methylase system may be present in G. violaceus.

Effect of CO2 concentrations on the activity of photosynthetic CO2 fixation and extracelluar carbonic anhydrase in the marine diatom Skeletonema costatum

The growth and activity of photosynthetic CO2 uptake and extracellular carbonic anhydrase (CA(ext)) of the marine diatom Skeletonema costatum were investigated while cultured at different levels of CO2 in order to see its physiological response to different CO2 concentrations under either a low (30 mumol . m(-2) . s(-1)) or high (210 mumol . m(-2) . s(-1)) irradiance. The changes in CO2 concentrations (4-31 mumol/L) affected the growth and net photosynthesis to a greater extent under the low than under the high light regime. CAext was detected in the cells grown at 4 mumol/L CO2 but not at 31 and 12 mumol/L CO2, with its activity being about 2.5-fold higher at the high than at the low irradiance. Photosynthetic CO2 affinity (1/K-1/2(CO2)) of the cells decreased with increased CO2 concentrations in culture. The cells cultured under the high-light show significantly higher photosynthetic CO2 affinity than those grown at the low-light level. It is concluded that the regulations of CA(ext) activity and photosynthetic CO2 affinity are dependent not only on CO2 concentration but also on light availability, and that the development of higher CA(ext) activity and CO2 affinity under higher light level could sufficiently support the photosynthetic demand for CO2 even at low level of CO2.

Salt tolerance of Microcoleus vaginatus Gom., a cyanobacterium isolated from desert algal crust, was enhanced by exogenous carbohydrates

Microcoleus vaginatus isolated from a desert algal crust of Shapotou was cultured in BG-11 medium containing 0.2mol l(-1) NaCl or 0.2mol l(-1) NaCl plus 100mg l(-1) sucrose, extracellular polymeric substances (EPS) or hot water-soluble polysaccharides (HWP), respectively. Photosynthetic oxygen evolution rates, photosystem 11 activity (Fv/Fm) and dark respiration of NaCl-stressed cells were enhanced significantly by the added sucrose or EPS under salt stress conditions (0.2mol l(-1) NaCl). Compared with cells treated with salt alone, sodium contents in cells reduced significantly; the content of cellular total carbohydrate did not change, and intracellular sucrose, water-soluble sugar increased significantly following the addition of exogenous carbohydrates. Sucrose synthase (SS) activity of NaCl-stressed cells increased following the addition of sucrose, and sucrose phosphate synthase (SPS) activity of NaCl-stressed cells increased following the addition of exogenous sucrose, EPS or HWP compared with cells stressed with NaCl only. The results suggested that the extruded EPS might be re-absorbed by cells of M. vaginatus as carbon source, they could increase salt tolerance of M. vaginatus through the changes of carbohydrate metabolism and the selective uptake of sodium ions. (C) 2003 Elsevier Science Ltd. All rights reserved.

pkn22 (alr2502) encoding a putative Ser/Thr kinase in the cyanobacterium Anabaena sp PCC 7120 is induced by both iron starvation and oxidative stress and regulates the expression of isiA

In cyanobacteria, the isiA gene is required for cell adaptation to oxidative damage caused by the absence of iron. We show here that a putative Ser/Thr kinase gene, pkn22 (alr2052), is activated by iron deficiency and oxidative damage in Anabaena sp. PCC 7120. A pkn22 insertion mutant is unable to grow when iron is limiting. pkn22 regulates the expression of isiA (encoding CP43') but not of isiB (encoding flavodoxin) and psbC (CP43). Fluorescence measurement at 77 K reveals the absence of the typical signature of CP43' associated with photosystem I in the mutant under iron-limiting conditions. We propose that Pkn22 is required for the function of isiA/CP43' and constitutes a regulatory element necessary for stress response. (C) 2003 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Marine species of Ainudrilus and Heterodrilus (Oligochaeta : Tubificidae : Rhyacodrilinae) from Hainan Island in southern China

Six species of Rhyacodrilinae (Oligochaeta: Tubificidae) are reported from intertidal and shallow water subtidal habitats around Hainan Island in southern China. Four species are new to science: Ainudrilus pauciseta n. sp., Heterodrilus chenianus n. sp., Heterodrilus nudus n. sp., and Heterodrilus uniformis n. sp. Japanese material of Ainudrilus lutulentus (Erseus, 1984) is also briefly described. Hitherto, 27 species belonging to Tubificidae have been recorded from Hainan.

Optimization of growth and fatty acid composition of a unicellular marine picoplankton, Nannochloropsis sp., with enriched carbon sources

A unicellular marine picoplankton, Nannochloropsis sp., was grown under CO2-enriched photoautotrophic or/and acetate-added mixotrophic conditions. Photoautotrophic conditions with enriched CO2 of 2800 mul CO2 l(-1) and aeration gave the highest biomass yield (634 mg dry wt l(-1)), the highest total lipid content (9% of dry wt), total fatty acids (64 mg g(-1) dry wt), polyunsaturated fatty acids (35% total fatty acids) and eicosapentaenoic acid (EPA, 20:5omega3) (16 mg g(-1) dry wt or 25% of total fatty acids). Mixotrophic cultures gave a greater protein content but less carbohydrates. Adding sodium acetate (2 mM) decreased the amounts of the total fatty acids and EPA. Elevation of CO2 in photoautotrophic culture thus enhances growth and raises the production of EPA in Nannochloropsis sp.

Effect of desert soil algae on the stabilization of fine sands

Four filamentous cyanobacteria, Microcoleus vaginatus, Phormidium tenue, Scytonema javanicum (Kutz.) and Nostoc sp., and a single-celled green alga, Desmococcus olivaceus, all isolated from Shapotou (Ningxia Hui Autonomous Region of China), were batch cultured and inoculated onto unconsolidated sand in greenhouse and field experiments. Their ability to reduce wind erosion in sands was quantified by using a wind tunnel laboratory. The major factors related to cohesion of algal crusts, such as biomass, species, species combinations, bioactivity, niche, growth phase of algae, moisture, thickness of the crusts, dust accretion (including dust content and manner of dust added) and other cryptogams (lichens, fungi and mosses) were studied. The best of the five species were M. vaginatus and P. tenue, while the best mix was a blend of 80% M. vaginatus and 5% each of P. tenue, S. javanicum, Nostoc sp. and D. olivaceus. The threshold friction velocity was significantly increased by the presence of all of the cyanobacterial species, while the threshold impact velocity was notably increased only by the filamentous species. Thick crusts were less easily eroded than thin crusts, while biomass was more effective than thickness. Dust was incorporated best into Microcoleus crust when added in small amounts over time, and appeared to increase growth of the cyanobacterium as well as strengthen the cohesion of the crust. Microbial crust cohesion was mainly attributed to algal aggregation, while lichens, fungi and mosses affected more the soil structure and physico-chemical properties.

Characterisation of a pathogenic virus isolated from marine threadfin fish (Eleutheronema tetradactylus) during a disease outbreak

An unknown virus was isolated from massive mortality of cultured threadfin (Eleutheronema tetradactylus) fingerlings. The virus replicated in BF-2 fish cell line and produced a plaque-like cytopathic effect. Electron micrographs revealed non-enveloped, icosahedral particles approximately 70-80 nm in diameter composed of a double capsid layer. Viroplasms and subviral particles approximately 30 run in diameter and complete particles of 70 nm in diameter were also observed in the infected BF-2 tissue culture cells. The virus was resistant upon pH 3 to 11 and ether treatment. It is also stable to heat treatment (3 h at 56 T). Replication was not inhibited by 5-iododeoxyuridine (5-IUdR). Acridine orange stain revealed typical reovirus-like cytoplasmic inclusion bodies. Electrophoresis of purified virus revealed 11 segments of double-stranded RNA and five major structural polypeptides of approximately 136, 132, 71, 41 and 33 kDa. Based on these findings, the virus isolated was identified to belong to the genus Aquareovirus and was designated as threadfin reovirus. This virus differed from a majority of other aquareovirus by its increase in virus infectivity upon exposure to various treatments such as high and low pH, heat (56 degreesC), ether and 5-IUdR. The RNA and virion protein banding pattern of the threadfin reovirus was shown to differ from another Asian isolate, the grass carp hemorrhage reovirus (GCV). Artificial injection of the threadfin reovirus into threadfin fingerlings resulted in complete mortality, whereas sea bass (Lates calcarifer) fingerlings infected via bath route showed severe mortality within a week after exposure. These results indicate that the threadfin virus is another pathogenic Asian aquareovirus isolate that could cross-infect into another marine fish, the sea bass. (C) 2002 Elsevier Science B.V. All rights reserved.