Fluorescence parameters of marine plankton algae at the assimilation of organic nitrogen

fluorescence parameters of marine plankton algae Pseudo-nitzschis delicatissima, Thalassiosira weissflogii, and Tetraselmis viridis were estimated after the addition of organic (urea and glycine) and inorganic (nitrate and ammonia) nitrogen to nitrogen-limited cultures acclimated to limited and saturated irradiance. The photochemical efficiency of photosystem 2, the maximum relative electron transport, and the light saturation index increased in the algae assimilating organic nitrogen. The dynamics of parameters depended species specifically on the nitrogen source and irradiance. The ecological role of organic nitrogen in the seasonal dynamics and vertical distribution of phytoplankton is discussed.     

ASSAY OPTIMIZATION AND REGULATION OF UREASE ACTIVITY IN TWO MARINE DIATOMS

An in vitro urease enzyme assay was developed for the marine diatoms Thalassiosira pseudonana Hasle et Heimdal (clone 3H) and T. weissflogii (Grunow) Fryxell et Hasle (clone Actin). This assay involves the colorimetric measurement of ammonium following the hydrolysis of urea in crude cell homogenates and it is the first assay to account for the rate of nitrogen assimilation in both species grown on urea as the sole nitrogen source. Urease activity was found to be present regardless of nitrogen source, although activities showed distinctly different patterns depending on the species examined and form of nitrogen supplied. Under nitrogen-replete conditions, urease activity in T. pseudonana was present constitutively when grown on NH₄⁺ and upregulated when grown on NO₃⁻ or urea. In nitrogen-replete T. weissflogii , urease activity was present at high constitutive levels regardless of the nitrogen source and showed no upregulation. Nitrogen starvation did not upregulate activity in either species.     

Did sulfate availability facilitate the evolutionary expansion of chlorophyll a+c phytoplankton in the oceans?

During the Mesozoic Era, dinoflagellates, coccolithophorids and diatoms became prominent primary producers in the oceans, succeeding an earlier biota in which green algae and cyanobacteria had been proportionally more abundant. This transition occurred during an interval marked by increased sulfate concentration in seawater. To test whether increasing sulfate availability facilitated the evolutionary transition in marine phytoplankton, the cyanobacterium Synechococcus sp., the green alga Tetraselmis suecica and three algae containing chlorophyll a+c (the diatom Thalassiosira weissflogii, the dinoflagellate Protoceratium reticulatum and the coccolithophorid Emiliania huxleyi) were grown in media containing 1, 5, 10, 20, or 30 mm SO(4) (2-) . The cyanobacterium and the green alga showed no growth response to varying [SO(4) (2-) ]. By contrast, the three chlorophyll a+c algae showed improved growth with higher [SO(4) (2-) ], but only up to 10 mm. The chlorophyll a+c algae, but not the green alga or cyanobacterium, also showed lower C:S with higher [SO(4) (2-) ]. When the same experiment was repeated in the presence of a ciliate predator (Euplotes sp.), T. suecica and T. weissflogii increased their specific growth rate in most treatments, whereas the growth rate of Synechococcus sp. was not affected or decreased in the presence of grazers. In a third experiment, T. suecica, T. weissflogii, P. reticulatum and Synechococcus sp. were grown in conditions approximating modern, earlier Paleozoic and Proterozoic seawater. In these treatments, sulfate availability, nitrogen source, metal availability and Pco(2) varied. Monospecific cultures exhibited their highest growth rates in the Proterozoic treatment. In mixed culture, T. weissflogii outgrew other species in modern seawater and T.suecica outgrew the others in Paleozoic water. Synechococcus sp. grew best in Proterozoic seawater, but did not outgrow eukaryotic species in any treatment. Collectively, our results suggest that secular increase in seawater [SO(4) (2-) ] may have facilitated the evolutionary expansion of chlorophyll a+c phytoplankton, but probably not to the exclusion of other biological and environmental factors.     

Fluorescence parameters of White Sea phytoplankton under different nitrogen sources

We investigated dependence of fluorescence parameters and phytoplankton biomass on the nitrogen source and irradiance in enriched flask studies with White Sea plankton from August-September 2007. Phytoplankton was exposed in situ for 18 d with addition of 180 μM/L of nitrogen in the forms of nitrate, urea, ammonia, and glycine under two levels of irradiance. Maximum quantum efficiency of PS2 (F_v/F_m) was determined in the samples adapted to darkness. Rapid light curves were obtained for each sample with the sequential increase of light intensity (8 levels). The maximal relative electron transport rate (rETR_max), the maximum light utilization coefficient (α), and the nonphotochemical quenching (NPQ) were calculated. The phytoplankton abundance increased on nitrogen addition, and the photosynthetic parameters changed. The values F_v/F_m reached 0.64–0.71, which indicated a good physiological state of algae and lack of nitrogen limitation. The dynamics of rETR_max and NPQ depended of the nitrogen source and irradiance, while α did not depend on nitrogen addition.     

Efficiency assessment of the one-step production of astaxanthin by the microalga Haematococcus pluvialis

Continuous cultivation of Haematococcus pluvialis under moderate nitrogen limitation represents a straightforward strategy, alternative to the classical two-stage approach, for astaxanthin production by this microalga. Performance of the one-step system has now been validated for more than 40 combinations of dilution rate, nitrate concentration in the feed medium, and incident irradiance, steady state conditions being achieved and maintained in all instances. Specific nitrate input and average irradiance were decisive parameters in determining astaxanthin content of the biomass, as well as productivity of the system. The growth rate of the continuous photoautotrophic cultures was a hyperbolic function of average irradiance. As long as specific nitrate input was above the threshold value of 2.7 mmol/g day, cells performed green and astaxanthin was present at basal levels only. Below the threshold value, under moderate nitrogen limitation conditions, astaxanthin accumulated to reach cellular levels of up to 1.1% of the dry biomass. Increasing irradiance resulted in enhancement of astaxanthin accumulation when nitrogen input was limiting, but never under nitrogen sufficiency. Mean daily productivity values of 20.8 +/- 2.8 mg astaxanthin/L day (1.9 +/- 0.3 g dry biomass/L day) were consistently achieved for a specific nitrate input of about 0.8 mmol/g day and an average irradiance range of 77-110 microE/m(2) s. Models relating growth rate and astaxanthin accumulation with both average irradiance and specific nitrate input fitted accurately experimental data. Simulations provided support to the contention of achieving efficient production of the carotenoid through convenient adjustment of the determining parameters, and yielded productivity estimates for the one-step system higher than 60 mg astaxanthin/L day. The demonstrated capabilities of this production system, as well as its product quality, made it a real alternative to the current two-stage system for the production of astaxanthin-rich biomass.     

Nitrogen Starvation and the Regulation of Glutamine Synthetase in Agmenellum quadruplicatum

The level of glutamine synthetase activity in Agmenellum quadruplicatum strain PR-6 was dependent on the nitrogen source used for growth and on the nutritional status of the cells. During exponential growth, glutamine synthetase activity was low in cells grown on ammonia, urea, or nitrate. During the transition from nitrogen replete to nitrogen starved growth, glutamine synthetase activity began to rise. With ammonia as a nitrogen source, glutamine synthetase activity as determined in whole cells increased from 1 nanomole per minute per milliliter during exponential growth to 22 nanomoles per minute per milliliter during severe nitrogen starvation. In cells grown on nitrate the increase was from 5 to 39 nanomoles per minute per milliliter, and in cells grown on urea the increase was from 4 to 31 nanomoles per minute per milliliter.     

不同氮源对蛋白核小球藻生长、色素和中性脂肪积累的影响

单细胞微藻在生长发育过程中,所积累的中性脂肪具有潜在的生物燃料价值。不同氮源对藻类的生长具有显著影响。研究了氨态氮、尿素氮和硝酸态氮对蛋白核小球藻生长、色素和中性脂肪积累的影响。结果显示,不同氮源对培养液pH有显著影响,以氨态氮为氮源导致培养液pH降低,而硝酸态氮导致培养液pH升高,培养液pH的波动可被添加的Hepes所稳定,并促进以氨态氮为氮源的蛋白核小球藻的生长。尿素氮对蛋白核小球藻生长、色素积累的效果优于氨态氮和硝酸态氮,硝酸态氮在中性脂肪积累上优于尿素氮和氨态氮,添加Hepes对氮饥饿后蛋白核小球藻的中性脂肪积累无显著影响。     

Effect of nitrogenous resource on growth,biochemical composition and ultrastructure of Isochrysis galbana (Isochrysidales,Haptophyta)

The nitrogenous resource used to promote algal growth has cost implications for mass culture processes. The present study therefore aimed to determine the effect of different nitrogenous resources (nitrate, ammonium and urea) on various performance parameters (growth, final cell yield, pigmentation, lipid yield and cellular and sub‐cellular characteristics) in Isochrysis galbana. Growth rate was unaffected by nitrogenous resource, but the final cellular yield in the nitrate and urea treatments far exceeded that evident in the ammonium treatments. The reduced cell yield in ammonium treatments and the earlier onset of the stationary phase was brought about by nitrogen‐starvation due to an increase in pH and resultant ammonia volatilization. This starvation initiated an early onset of lipid accumulation, chlorophyll depletion and an increase in the carotenoid to chlorophyll ratio relative to the other nitrogen (N) source treatments. Hence, in spite of being potentially the preferred source of N by algae (due to its reduced state), ammonium‐nitrogen is undesirable for mass culture. The performance parameters of Isochrysis grown in urea (an organic N source) and nitrate (an inorganic N source) were similar, but lipid accrued earlier in cells grown in medium supplemented with urea. This is advantageous for lipid acquisition for the production of biodiesel since it would reduce the duration of photobioreactor runs. Urea is easily available and considerably cheaper than all the other N sources tested and is thus recommended as the nitrogenous resource for large‐scale culture of I. galbana for biodiesel production.     

The Influence of Urea and Other Nitrogen Sources on Growth Rate of Scots Pine Seedlings

The growth rate and water content of urea-fed seedlings of Pinus silvestris L. were compared with those of nitrate-and ammonium-fed seedlings grown in continuously renewed nutrient solutions, in which the hydrolysis of urea to ammonia and carbon dioxide was minimized. The growth rate of seedlings grown in an ammonium nutrient solution, in an urea nutrient solution and in a nitrate nutrient solution was about 90 per cent, 75 per cent and 60 per cent, respectively, of that of seedlings grown in a mixture of ammonium and nitrate. Seedlings with urea as the sole nitrogen source developed very severe chlorosis of the needles, the old roots were dark-coloured, the whole root system was very fragile, and the lateral roots of the third order were missing. Urea-grown seedlings had the highest nitrogen contents, closely followed by the ammonium and the ammonium + nitrate seedlings. The lowest nitrogen level was in nitrate seedlings. The low growth rate and the chlorosis of urea-fed seedlings were suggested to be the result of a hydrolysis of urea inside the root, causing an increase in pH and an accumulation of ammonia in the root.     

Differential Effects of Nitrogen Limitation on Photosynthetic Efficiency of Photosystems I and II in Microalgae

The effects of nitrogen starvation on photosynthetic efficiency were examined in three unicellular algae by measuring changes in the quantum yield of fluorescence with a pump-and-probe method and thermal efficiency (i.e. the percentage of trapped energy stored photochemically) with a pulsed photoacoustic method together with the inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethylurea to distinguish photosystems I and II (PSI and PSII). Measured at 620 nm, maximum thermal efficiency for both photosystems was 32% for the diatom Thalassiosira weissflogii (PSII:PSI ratio of 2:1), 39% for the green alga Dunaliella tertiolecta (PSII:PSI ratio of 1:1), and 29% for the cyanobacterium Synechococcus sp. PCC 7002 (PSII:PSI ratio of 1:2). Nitrogen starvation decreased total thermal efficiency by 56% for T. weissflogii and by 26% for D. tertiolecta but caused no change in Synechococcus. Decreases in the number of active PSII reaction centers (inferred from changes in variable fluorescence) were larger: 86% (T. weissflogii), 65% (D. tertiolecta), and 65% (Synechococcus). The selective inactivation of PSII under nitrogen starvation was confirmed by independent measurements of active PSII using oxygen flash yields and active PSI using P700 reduction. Relatively high thermal efficiencies were measured in all three species in the presence of the PSII inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethylurea, suggesting the potential for significant cyclic electron flow around PSI. Fluorescence or photoacoustic data agreed well; in T. weissflogii, the functional cross-sectional area of PSII at 620 nm was estimated to be the same using both methods (approximately 1.8 x 102 A2). The effects of nitrogen starvation occur mainly in PSII and are well represented by variable fluorescence measurements.     

Studies on the purification of wastewater from the nitrogen fertilizer industry by intensive algal cultures. I. Growth of Chlorella vulgaris in wastes.

The possibility of growth of intensive cultures of Chlorella vulgaris on industrial wastewater from nitrogen fertizer plant containing ammonia, urea and nitrate was investigated. Good growth of algae was obtained when the waste was enriched with phosphorus and inoculum contained a high number of cells. The optimal pH for the culture was 7.0--8.0. The main factor limiting growth of algae on wastes on the concentration of ammonia nitrogen. Chlorella vulgaris grows quite well in wastes containing 600 mg NH4-N/l but is inhibited at concentration about 100 mg NH4-N/l.     

A microcosm study of nitrogen utilization in the Great Salt Lake,Utah

Microcosms were used to study the effects of two inorganic nitrogen sources (ammonia and nitrate) and two organic nitrogen sources (urea and glutamic acid) on the growth of algae and bacteria found in the Great Salt Lake, Utah. Ammonia, nitrate and urea stimulated bacterial growth indirectly through increased algal production of unknown organic substances. Glutamic acid, representing readily available organic carbon and nitrogen, stimulated the bacteria directly. No nitrification was observed in the microcosms although nitrite was found when the microcosms were supplemented with nitrate. Lake sediment contained a number of anaerobic bacteria producing hydrogen sulfide, methane and other gases. Production of these gases was stimulated in the columns with high algal and bacterial activity.     

Fenology and life cycle of Gracilariopsis tenuifions (Gracilariaceae) at Sucre, Venezuela

The aim of this work was to determine the frequency of different reproductive phases and to induce the germination of spores from tetrasporic and cystocarpic Gracilariopsis tenuifrons from Chacopata and La Pe?a, Venezuela, under controlled laboratory conditions (temperature 22 +/- 1 degree C, 12L:12D photoperiod, salinity of 36 +/-1 PSU and irradiance of 269 microE m(-2) s(-1)). Tetrasporic individuals dominated numerically over gametophitic individuals. The proportion of vegetative algae was very low. Only cystocarpic algae were collected and the spermatangia were absent. Sporulation, germination and formation of algae suggest that they present a Polysiphonia-type life cycle. Algae with reproductive structures were not obtained in the laboratory,     

The utilization of organic nitrogen for growth of algae: physiological aspects

The ability of 27 algae belonging to 11 taxonomic divisions to grow at the expense of organic nitrogen was tested in axenic culture. Experiments were carried out in buffered media: morpholine propanesulphonic acid (MOPS) was used successfully for all fresh water strains except diatoms. It was not used as a nitrogen source by any strain examined but served as a good source of sulphur for four strains. The range of substrates used was extensive and included amino acids, urea, acetamide, urate and some nucleosides. Growth rates varied widely but growth yields were generally comparable to those attained with nitrate or ammonia except for substrates containing more than one utilizable nitrogen atom. Limited experiments were carried out in the dark, and it was found that a given substrate was equally suitable for both dark heterotrophic and photolithotrophic growth. Levels of chlorophyll a were measured during growth of two algae with nitrate, glycine and urate. With nitrate, levels of specific chlorophyll a declined some ten-fold during growth: with poor substrates supporting only slow growth, levels were more uniform but were only 10–15% of those attained during growth with good substrates. One strain grown with MOPS as sole source of sulphur produced cultures with levels of chlorophyll a about half those found in sulphate-grown cultures. Tolerance of sodium chloride was examined in a few strains and even putatively fresh water strains were found to be appreciably tolerant.     

Structure of the community of planktic algae of the white sea in relation to availability of oxidized or reduced forms of nitrogen and level of illumination

Phytoplankton sampled in Kandalaksha Bay of the White Sea in late June 2006 was exposed in situ for 14 days under two levels of illumination (E₁ > E₂) with the addition of 180 μM of nitrogen as nitrate, urea, ammonium, and glycine. Every 3 days, species composition, abundance, and biomass of algae were estimated. The maximum biomass (B_max) of phytoplankton depended on nitrogen substrates and level of illumination. The differences in Bmax between phytoplankton assimilating different nitrogen substrates became more apparent under E₂. Phytoplankton assimilating nitrates, urea, and ammonium had higher B_max under E₁, but algae grown with glycine had higher Bmax under E₂. Although the values of B_max differed, the structures of all assemblages were similar except for that grown on ammonium under E₁. Competitive parameters of algae populations depended on the form of nitrogen and level of illumination in a species-specific manner. Comparison of the present data with results of similar experiments in 2005 revealed that structure of assemblages formed after nitrogen pulse additions depended not only on nitrogen forms and illumination level but also on initial (before pulse) structure of community and availability of nitrogen for algae.     

Light, temperature and nitrogen as interacting factors affecting diel vertical migrations of dinoflagellates in culture

Diel vertical migrations of the marine dinoflagellates Gonyaulaxpolyedra Stein and Ceratium furca (Ehr.) Clap, et Lachm. werefollowed in a laboratory tube (2.02 m x 0.25 m) under a 12:12hlight:dark cycle. The effects of temperature stratification,two levels of surface irradiance and nitrogen depletion on patternsof vertical migrations were examined. At temperatures between22–26°C with small temperature gradients, both speciesmigrated at a rate of 0.7 –1.0 m h^–1. Steeper thermoclines(ca. 0.8°C 0.1 m^–1) with temperatures below ca. 20°Ccaused a marked decrease in swimming speed which resulted inaccumulations of cells in these thermocline regions. Under conditionsof nutrient sufficiency both algae migrated into the surfacelayers at irradiance values of over 1000 µE m^–2s^–1. Increasing nitrogen depletion caused the downwardmigration of both algae to commence progressively earlier inthe day and before the end of the light period. The earlierdownward migrations enabled a more complete descent throughthe thermocline. Nitrogen depleted cells of Gonyaulax continuedto undertake vertical migrations but avoided high irradiancesthus forming subsurface maxima at irradiance levels close to150 µE m^–2 s^–1. Ceratium cells which exhaustedboth inorganic nitrogen and phosphorus ceased to migrate accompaniedby a large change in cellular fluorescence.     

Regulation of Growth at Steady-state Nitrogen Nutrition in Lettuce (Lactuca sativa L.): Interactive Effects of Nitrogen and Irradiance

The morphological and physiological adaptation of Lactuca sativaL. (‘Vegas’) to different irradiance levels andrates of nitrogen supply was analysed in such a way that effectsof irradiance were clearly distinguished from the effects ofnitrogen. Lettuce was grown in a glasshouse in aerated nutrientsolutions containing all necessary nutrients except nitrogen.Nitrogen was supplied in excess and at limiting rates in relationto plant growth to provide steady state nutrition. Shading theplants created the low irradiance level. The effects of nitrogensupply and irradiance on growth showed a marked interaction.Dry matter production decreased strongly with decreasing nitrogensupply at high irradiance, but decreased only slightly at lowirradiance. Nitrogen had no effect on radiation use efficiencyexcept for the lowest nitrogen treatment at high irradiance.The effect of nitrogen on growth was mainly mediated by itseffect on leaf area development and hence on light interception.Decreases in leaf area were due to decreases in specific leafarea and dry matter partitioning towards the leaves, while thedecrease in specific leaf area was the result of an increasein leaf dry matter percentage at low nitrogen supply. Dry matterand nitrogen partitioning, and nitrate concentration were closelyrelated to plant nitrogen concentration. Irradiance did notaffect these relationships. Irradiance influenced partitioningonly indirectly by affecting plant nitrogen concentration. Thedemand for organic nitrogen per unit leaf area was lower atlower irradiance. Organic nitrogen per unit leaf area appearedto be adjusted to the irradiance level, independently of thenitrogen supply, suggesting priority of nitrogen usage in photosynthesis.Copyright 2000 Annals of Botany Company Lactuca sativa L., lettuce, growth, irradiance, leaf area, nitrogen, radiation use efficiency, partitioning     

Distribution of Nitrogen during Growth of Sunflower (Helianthus annuus L.)

The accumulation, distribution and redistribution of dry matterand nitrogen is described for Helianthus annuus L. cv. Hysun21 grown on 6 mM urea in glasshouse culture. Seed dry matterand nitrogen were transferred to seedlings with net efficienciesof 40 and 86 per cent respectively. At flowering, the stem hadmost of the plant's dry matter and the leaves most of its nitrogen.About 35 per cent of the plant's nitrogen accumulated afterthree-row anthesis. The amount of protein in vegetative parts,especially leaves, declined after flowering. Concentrationsof free amino compounds also decreased during growth. Matureseeds had 38 per cent of the total plant dry weight and 68 percent of the total nitrogen. Seeds acquired 33 per cent of theirdry matter and nitrogen from redistribution from above-groundplant parts. The stem was most important for storage of carbohydrate,leaves the most important for nitrogen. Over 50 per cent ofthe nitrogen in the stem and leaves was redistributed. Plantsthat received 6 mM nitrate accumulated more dry matter thanurea-grown plants. Seeds from nitrate-grown plants were heavier(58 mg) than those of urea-grown plants (46 mg), and their percentageoil was greater (50 and 41 respectively). The amount of nitrogenper seed was the same. Little or no urea was detected in xylem sap of plants suppliedwith 5 mM urea, but it was detected in sap of plants which received25 mM. Concentrations of urea and amino compounds in the sapdecreased up the stem. Plants supplied with nitrate had mostof the nitrogen in xylem sap as NO₂^–, suggesting littlenitrate reduction in roots. Plants grown on 6 mM nitrate andchanged to high levels of urea-nitrogen for 14 days still hadhigh levels of nitrate; little nitrate remained in plants receivinglow levels of urea. When urea is applied in irrigation waterto field-grown sunflower, the nitrogen is subsequently takenup as nitrate due to rapid nitrogen transformations in the soil. Helianthus annuus L., sunflower, urea, nitrate, nitrogen transport, xylem sap, nitrogen accumulation nitrogen distribution