UNCOUPLING OF SILICON COMPARED WITH CARBON AND NITROGEN METABOLISMS AND THE ROLE OF THE CELL CYCLE IN CONTINUOUS CULTURES OF THALASSIOSIRA PSEUDONANA (BACILLARIOPHYCEAE) UNDER LIGHT,NITROGEN, AND PHOSPHORUS CONTROL1

The elemental composition and the cell cycle stages of the marine diatom Thalassiosira pseudonana Hasle and Heimdal were studied in continuous cultures over a range of different light‐ (E), nitrogen‐ (N), and phosphorus‐ (P) limited growth rates. In all growth conditions investigated, the decrease in the growth rate was linked with a higher relative contribution of the G2+M phase. The other phases of the cell cycle, G1 and S, showed different patterns, depending on the type of limitation. All experiments showed a highly significant increase in the amount of biogenic silica per cell and per cell surface with decreasing growth rates. At low growth rates, the G2+M elongation allowed an increase of the silicification of the cells. This pattern could be explained by the major uptake of silicon during the G2+M phase and by the independence of this process on the requirements of the other elements. This was illustrated by the elemental ratios Si/C and Si/N that increased from 2‐ to 6‐fold, depending of the type of limitation, whereas the C/N ratio decreased by 10% (E limitation) or increased by 50% (P limitation). The variations of the ratios clearly demonstrate the uncoupling of the Si metabolism compared with the C and N metabolisms. This uncoupling enabled us to explain that in any of the growth condition investigated, the silicification of the cells increased at low growth rates, whereas carbon and nitrogen cellular content are differently regulated, depending of the growth conditions.     

EFFECT OF NUTRIENT AVAILABILITY ON THE BIOCHEMICAL AND ELEMENTAL STOICHIOMETRY IN THE FRESHWATER DIATOM STEPHANODISCUS MINUTULUS (BACILLARIOPHYCEAE)*

The objective of this study was to examine the differences in the biochemical and elemental stoichiometry of a freshwater centric diatom, Stephanodiscus minutulus (Grun.), under various nutrient regimes. Stephanodiscus minutulus was grown at μ_max or 22% of μ_max under limitation by silicon, nitrogen, or phosphorus. Cell sizes for nutrient‐limited cultures were significantly smaller than the non‐limited cell sizes, with N‐limited cells being significantly smaller than all other treatments. Compared with the nutrient‐replete treatment, both carbohydrates and lipids increased in Si‐ and P‐limited cells, whereas carbohydrates increased but proteins decreased in N‐limited cells. All of the growth‐limited cells showed an increase of carbohydrate and triglyceride, and a decrease of cell size and polar lipids as a percentage of total lipids. The non‐limited cells also had a significantly higher chl a concentration and galactolipids as a percentage of total lipids than any of the limited treatments, and the low‐Si and low‐P cells had significantly higher values than the low‐N cells. The particulate C concentrations showed significant differences between treatments, with the Si‐ and P‐limited treatments being significantly higher than the N‐ and non‐limited treatments. Particulate Si did not show a strong relationship with any of the parameters measured, and it was the only parameter with no differences between treatments. The low‐Si cells had a significantly higher P content (about two times more) than any other treatment, presumably owing to the luxury consumption of P, and a correspondingly high phospholipid concentration. The elemental data showed that S. minutulus had a high P demand with low optimum N:P (4) and Si:P (10) ratios and a C:N:P ratio of 109:16:2.3. The particulate C showed a positive relationship with POM (r = 0.93), dry weight (r = 0.88), lipid (r = 0.87) and protein (r = 0.84, all P < 0.0001). Particulate N showed a positive relationship with galactolipids (r = 0.95), protein (r = 0.90), dry weight (r = 0.78), lipid (r = 0.75), and cell volume (r = 0.64, all P < 0.0001). It is evident that nutrient limitation in the freshwater diatom S. minutulus has pronounced effects on its biochemical and elemental stoichiometry.     

Increase in Si:N drawdown ratio due to resting spore formation by spring bloom-forming diatoms under Fe- and N-limited conditions in the Oyashio region

Resting spore formation and Si:N drawdown ratios were investigated under iron (Fe)- and nitrogen (N)-limited conditions using a unialgal culture of Thalassiosira nordenskioeldii and natural phytoplankton assemblages during the spring bloom in the Oyashio region. In the unialgal culture of T. nordenskioeldii, 20% and 100% of the cells formed resting spores under Fe- and N-limited conditions, respectively. The Si:N drawdown ratios were 2- and 14-fold higher in Fe- and N-limited conditions, respectively, compared to Fe- and N-sufficient conditions. At the start of the natural phytoplankton incubation, 18 among 47 identified diatom species were known resting spore-forming species. Approximately 15 common diatom species formed resting spores under Fe- and N-limited conditions. During the natural phytoplankton incubation, the percentage of the resting spores increased with time under both Fe- and N-limited conditions, reaching 25% and 40% of total diatom abundance, respectively. The Si:N drawdown ratios significantly increased with an increase in the contribution of resting spores in both the unialgal culture and natural phytoplankton incubations. These results suggest that if the bloom dominated by neritic, resting spore-forming diatom species decline by either Fe- or N-depletion, Si may be utilized preferentially to N in the upper mixed layer due to the formation of heavily silicified resting spores.     

EFFECTS OF Si:P CONCENTRATION RATIOS AND NUTRIENT LIMITATION ON THE CELLULAR COMPOSITION AND MORPHOLOGY OF ASTERIONELLA FORMOSA (BACILLARIOPHYCEAE)1

The freshwater diatom Asterionella formosa Haas. was grown in semicontinuous culture at 20°C under continuous cool-white fluorescent light of ca. 20 μEin · m^?2· s ·^?1 in a medium containing Si: P in various concentration ratios. The cell quotas of P and Si changed in relation to the available concentrations of P and Si at constant μ= 0.11 and 0.16 d^?1. Under Si-limitation, the P cell quota increased by over an order of magnitude as the influent [Si:P] decreased. The Si cell quota increased with increase in [Si] in the influent medium, and it increased as [P] increased at a specific [Si]. Under P-limitation, the P cell quotas were fairly constant and low; the Si cell quotas were relatively high and decreased slightly as influent [P] and [Si] increased. Asterionella stored up to 28 times more P and 2 times more Si than needed. The number of Asterionella cells per colony varied as a function of the influent [Si:P] and nutrient limitation being usually less than or equal to 6 when P-limited, and greater than 10 when Si-limited.     

The relationship between carbon and biovolume in marine microbial mesocosms under different nutrient regimes

Nutrient manipulation experiments were conducted on a natural planktonic community in outdoor mesocosms. Inorganic nitrogen (N) and silicon (Si) were added to achieve N:Si ratios of 1:1 and 4:1. Total particulate carbon (PC) biomass of the microbial assemblage was determined by elemental analysis. Cell volume measurements by microscope on individual components of the community (bacteria, diatoms, photosynthetic nanoflagellates, heterotrophic nanoflagellates, dinoflagellates and ciliates) were also made. We applied published C:volume relationships to determine the volume estimated C content (CBV) of these microbial groups and hence of the total assemblage. The total CBV and total PC were compared to test the applicability of C:volume relationships under different nutrient regimes both before and after nutrient exhaustion. For initial N:Si ratios of 1:1, prior to nutrient exhaustion, the relationship between CBV and PC was linear with a gradient of approximately 1, (0.99?±?0.06), indicating that the published C:volume relationships accurately predicted the C content of the microbial assemblage. For N:Si ratios of 4:1, a linear relationship was again evident between CBV and PC (slope: 1.36?±?0.08). However, statistical comparison using a general linear model indicated that the gradient of this relationship differed significantly from that when the N:Si ratio was 1:1, and hence CBV overestimated elemental C. For both N:Si ratios, subsequent to nutrient exhaustion (N or Si), and hence when the diatom fraction of the microbial assemblage was in yield-limited post-exponential phase, the two measures of biomass were not well correlated. This indicated that measured cytoplasmic cell volume was a poor indicator of C biomass within the microbial assemblage in nutrient-deplete conditions.     

Effects of temperature,irradiance, and daylength on the marine diatom leptocylindrus danicus cleve. I. Photosynthesis and cellular composition

Rates of ¹⁴C uptake and cellular composition of C, N, and Chl a in the marine diatom Leptocylindrus danicus Cleve were measured in axenic batch culture under 49 combinations of temperature (5, 10, 15, 20 °C), daylength (15: 9, 12: 12, 9: 15 LD), and irradiance (at least four irradiances per daylength). ¹⁴C uptake exhibited a temperature-dependent daylength effect. Similar P-I curves characterized cells grown under 15: 9 and 12: 12 LD; P_max values were 17.2, 11.2, 4.3, and 1.8 pg C. pg Chl a^?1. h^?1 at 20, 15, 10, and 5°C, respectively. Under 9:15 LD at 20 and 15°C, the lightsaturated photosynthetic rate was ≈50% that in cells grown under longer daylengths. ¹⁴C uptake was independent of daylength at 10 and 5°C. The initial slope, a, of cells grown under long daylengths increased by five-fold between 5 and 20 °C. α values of cells grown under 9: 15 LD at 15 and 20 °C were depressed relative to longer daylengths. Chl a was inversely related to irradiance, and increased with temperature from 10 to 20 °C, whereas cell carbon and nitrogen showed a similar temperature dependence but was not influenced by irradiance or daylength. The C : N ratio and cell volume were independent of temperature, irradiance, and daylength. Both the C : Chl a and N : Chl a ratios increased with irradiance by greater amounts at lower temperatures.     

KINETICS OF SILICIC ACID UPTAKE AND RATES OF SILICA DISSOLUTION IN THE MARINE DIATOM THALASSIOSIRA PSEUDONANA1,2

Tracer techniques using the stable isotope ³⁰Si were used to measure rates of silicic acid uptake and silica dissolution in silicon replete and silicon depleted populations of 2 clones of the marine diatom Thalassiosira pseudonana Hasle & Heimdal. Uptake kinetics were describable using the Michaelis-Menten equation for enzyme kinetics, and no threshold concentration for uptake was evident. The maximum specific uptake rate of the estuarine clone 3H (0.062–0.092 · h^?1) was higher than that of the Sargasso Sea clone 13-1 (0.028–0.031 · h^?1), but half-saturation constants for uptake by the 2 clones were not measurably different (0.8–2.3 μM for 3H; 1.4–1.5 μM for 13-1). There was little or no light dependence of uptake in populations grown under optimal light conditions prior to the experiment. Exponentially growing populations released silicic acid to the medium by dissolution of cellular silica at rates ranging from 6.5 to 15% of the maximum uptake rate.     

STUDIES ON THE AUTECOLOGY OF THE MARINE DIATOM THALASSIOSIRA NORDENSKIÖLDIICLEVE. 1. THE INFLUENCE OF DAYLENGTH,LIGHT INTENSITY,AND TEMPERATURE ON GROWTH1

The marine diatom Thalassiosira nordenskiöldii Clave was grown at 48 different combinations of daylength (9:15, 12:12, 15:9 LD), light intensity (0.011, 0.027, 0.066, 0.100 ly/min [g cal/cm²/min]), and temperature (0, 5, 10, 15 C). Growth occurred at all combinations of light and temperature except at 15 C at the highest light level. Maximum growth (K = 1.8 doublings/day) occurred at 10 C under the 15:9 LD cycle. At 15 C the maximum rate was 1.7 doublings/day but occurred at the shortest day-length (9:15 LD). The maxima at 5 and 0 C were 1.32 and 0.67 doubling/day, respectively. At 0 C growth was similar over a wide range of light intensities (K = 0.6–0.65), with, maximum growth being attained at a much lower light intensity than at 5 C. Above 5 C there was a decrease in the light intensity at which maximum growth occurred and excessive light became inhibitory to growth. At 15 C the light intensity at which maximum growth occurred was greater with shorter day-lengths. The temperature optimum was 10 C at 15:9 and 15 C at 9:15 LD. The chlorophyll a content of the cells was greatest under low light intensities and short daylengths, while temperature had a variable effect. The response of Thalassiosira in the laboratory contrasts with, its apparent preference for low temperatures in nature (0–5 C). The experiments suggest that the termination of the bloom of Thalassiosira in Narragansett Bay and elsewhere is not solely temperature dependent.     

PRESENCE OF CIRCADIAN RHYTHMS IN THE LOCOMOTOR ACTIVITY OF A CAVE-DWELLING MILLIPEDE GLYPHIULUS CAVERNICOLUS SULU (CAMBALIDAE,SPIROSTREPTIDA)

The locomotor activity of the millipede Glyphiulus cavernicolus (Spirostreptida), which occupies the deeper recesses of a cave, was monitored in light-dark (LD) cycles (12h light and 12h darkness), constant darkness (DD), and constant light (LL) conditions. These millipedes live inside the cave and are apparently never exposed to any periodic factors of the environment such as light-dark, temperature, and humidity cycles. The activity of a considerable fraction of these millipedes was found to show circadian rhythm, which entrained to a 12:12 LD cycle with maximum activity during the dark phase of the LD cycle. Under constant darkness (DD), 56.5% of the millipedes (n = 23) showed circadian rhythms, with average free-running period of 25.7h ± 3.3h (mean ± SD, range 22.3h to 35.0h). The remaining 43.5% of the millipedes, however, did not show any clear-cut rhythm. Under DD conditions following an exposure to LD cycles, 66.7% (n = 9) showed faint circadian rhythm, with average free-running period of 24.0h ± 0.8h (mean ± SD, range 22.9h to 25.2h). Under constant light (LL) conditions, only 2 millipedes of 11 showed free-running rhythms, with average period length of 33.3h ± 1.3h. The results suggest that these cave-dwelling millipedes still possess the capacity to measure time and respond to light and dark situations. (Chronobiology International, 17(6), 757–765, 2000)     

Circadian Rhythms of Embryonic Development and Hatching in Fish: A Comparative Study of Zebrafish (Diurnal), Senegalese Sole (Nocturnal), and Somalian Cavefish (Blind)

During early development, most organisms display rhythmic physiological processes that are shaped by daily changes in their surrounding environment (i.e., light and temperature cycles). In fish, the effects of daily photocycles and their interaction with temperature during early developmental stages remain largely unexplored. We investigated the existence of circadian rhythms in embryonic development and hatching of three teleost species with different daily patterns of behavior: diurnal (zebrafish), nocturnal (Senegalese sole), and blind, not entrained by light (Somalian cavefish). To this end, fertilized eggs were exposed to three light regimes: 12 h of light: 12 h of darkness cycle (LD), continuous light (LL), or continuous darkness (DD); and three species-appropriate temperature treatments: 24°C, 28°C, or 32°C for zebrafish and cavefish and 18°C, 21°C, or 24°C for sole. The results pointed to the existence of daily rhythms of embryonic development and hatching synchronized to the LD cycle, with different acrophases, depending on the species: zebrafish embryos advanced their developmental stage during the light phase, whereas sole did so during the dark phase. In cavefish, embryogenesis occurred within 24 h post fertilization (hpf) at the same pace during day or night. The hatching rhythms appeared to be controlled by a clock mechanism that restricted or “gated” hatching to a particular time of day/night (window), so that embryos that reached a certain developmental state by that time hatch, whereas those that have not wait until the next available window. Under LL and DD conditions, hatching rhythms and the gating phenomenon persisted in cavefish, in zebrafish they split into ultradian bouts of hatching occurring at 12–18-h intervals, whereas in sole DD and LL produced a 24-h delay and advance, respectively. Hatching rates were best under the LD cycle and the reported optimal temperature for each species (95.2?±?2.7% of the zebrafish and 83.3?±?0.1% of the cavefish embryos hatched at 28°C, and 93.1?±?2.9% of the sole embryos hatched at 21°C). In summary, these results revealed that hatching rhythms in fish are endogenously driven by a time-keeping mechanism, so that the day and time of hatching are determined by the interplay between the developmental state (temperature-sensitive) and the circadian clock (temperature-compensated), with the particular phasing being determined by the diurnal/nocturnal behavior of the species. (Author correspondence: javisan@um.es)     

Diatom elemental and morphological changes in response to iron limitation: a brief review with potential paleoceanographic applications

Diatoms are a major group of phytoplankton that account for approximately 40% of the ocean carbon fixation and the vast majority of biogenic silica production through the construction of their cell walls (termed frustules). These frustules accumulate and are partially preserved in the ocean sediments. Diatom growth and nutrient utilization in high‐nitrate, low‐chlorophyll regions of the world’s oceans are mostly regulated by iron availability. Diatoms acclimate to iron limitation by decreasing cell size. The associated increase in surface area‐to‐volume ratio and decrease in diffusive boundary layer thickness may improve nutrient uptake kinetics. In parallel, cellular silicon (Si) contents are elevated in iron‐limited diatoms relative to nitrogen (N) and carbon (C). Variations in degree of silicification and nutritional requirements of iron‐limited diatoms have been hypothesized to account for higher cellular Si and/or lower cellular N and C, respectively. However, in some diatoms, frustule silicification does not significantly change when cells are iron‐limited. Instead, changes in the Si‐containing valve surface area relative to volume within these diatoms is hypothesized to be responsible for the variations in the cellular Si : N and Si : C ratios. In particular, some examined iron‐limited pennate diatoms have reduced widths relative to their lengths (i.e. lower length‐normalized widths, LNW) compared to iron‐replete cells. In the pennate diatom Fragilariopsis kerguelensis, the mean LNWs of valves preserved in sediments throughout the Southern Ocean (a well‐characterized iron‐limited region) is positively correlated with satellite‐derived, climatological net primary productivity in the overlying waters. Because of the specific morphological changes in pennate diatom frustules in response to iron availability, the valve morphometerics (e.g. LNWs) can potentially be used as a diagnostic tool for iron‐limited diatom growth and relative changes in the Si : N (and Si : C) ratios in extant diatom assemblages as well as those preserved in the sediments.     

Species specificity in photoperiodic control of nymphal development in four species of cricket from north-west China

Photoperiodic regulation of nymphal development was examined in four species of cricket collected in the Xinjiang‐Uygur Autonomy Region, China (approximately 43°N, 81–89°E). Fifty percent of individuals of Modicogryllus frontalis reared at 28°C reached adulthood in approximately 80 days in conditions of 11 h light : 13 h dark (LD 11:13) to 14:10, and in approximately 95 days under LD 15:9 to 16:8. Melanogryllus desertus started adult emergence earliest under LD 16:8 at 28°C, but some individuals required much longer to mature, and thus two peaks of adult emergence occurred at approximately 60 and 160 days after hatching. More individuals hatched during the late peak in LD 18:6 than in LD 16:8. The mean nymphal period was approximately 100 days in LD 11:13 to 14:10. Both species showed short‐day type photoperiodic responses, but Mo. frontalis developed faster than did Me. desertus. The latter occupied a wider range of habitat conditions and was more variable in life cycle than the former. Modicogryllus burdigalensis started adult emergence earliest in LD 16:8 at 28°C in the six photoperiods tested, most adults emerging within 60 days. The mean nymphal period was 80 days in LD 15:9, 135 days in LD 14:10 and 80–100 days in LD 11:13 to 13:11, showing an intermediate type of photoperiodic response. Acheta domesticus was a long‐day type species, and the proportion of delayed individuals increased with decreasing photoperiod. In the dry climate of Xinjiang‐Uygur, nymphal overwintering seems to be successful in all of the four different patterns of nymphal development.     

EFFECTS OF HARVEST STAGE AND LIGHT ON THE BIOCHEMICAL COMPOSITION OF THE DIATOM THALASSIOSIRA PSEUDONANA1

The marine diatom Thalassiosira pseudonana (Hustedt, clone 3H) Hasle and Heimdal was cultured under three different light regimes: 100 μmol photon · m^?2· s^?1 on 12:12 h light : dark (L:D) cycles; 50 μmol photon · m^?2· s^?2 on 24:0 h L:D; and 100 μmol photon · m^?2· s^?1 on 24:0 h L:D. It was harvested during logarithmic and stationary phases for analysis of biochemical composition. Across the different light regimes, protein (as % of organic weight) was highest in cells during logarithmic phase, whereas carbohydrate and lipid were highest during stationary phase. Carbohydrate concentrations were most affected by the different light regimes; cells grown under 12:12 h L:D contained 37–44% of the carbohydrate of cells grown under 24:0 h L:D. Cells in logarithmic phase had high proportions of polar lipids (79 to 89% of total lipid) and low triacylglycerol (≤10% of total lipid). Cells in stationary phase contained less polar lipid (48 to 57% of total lipid) and more triacylglycerol (22 to 45% of total lipid). The fatty acid composition of logarithmic phase cells grown under 24:0 h L:D were similar, but the 100 μmol photon · m^?2· s^?1 (12:12 h L:D) cells at the same stage contained a higher proportion of polyunsaturated fatty acids (PUFAs) and a lower proportion of saturated and monounsaturated fatty acids due to different levels of 16:0, 16:1(n-7), 16:4(n-1), 18:4(n-3), and 20:5(n-3). With the onset of stationary phase, cells grown at 100 μmol photon · m^?2· s^?1 (both 12:12 and 24:0 h L:D) increased in proportions of saturated and monounsaturated fatty adds and decreased in PUFAs. Concentrations (% organic or dry weight) of 14:0, 16:0, 16:1(n-7), 20:5(n-3), and 22:6(n-3) increased in cells of all cultures during stationary phase. The amino acid compositions of cells were similar irrespective of harvest stage and light regime. For mariculture, the recommended light regime for culturing T. pseudonana will depend on the nutritional requirements of the animal to which the alga is fed. For rapidly growing bivalve mollusc larvae, stationary-phase cultures grown under a 24:0 h L:D regime may provide more energy by virtue of their higher percentage of carbohydrate and high proportions and concentrations of energy-rich saturated fatty acids.     

STOICHIOMETRIC RESPONSES OF PHYTOPLANKTON SPECIES TO THE INTERACTIVE EFFECT OF NUTRIENT SUPPLY RATIOS AND GROWTH RATES1

Three species of phytoplankton, Rhodomonas sp., Phaeodactylum tricornutum Bohlin, and Isochrysis galbana Parke, were cultivated in semicontinuous culture to analyze the response of carbon (C):nitrogen (N):phosphorus (P) stoichiometry to the interactive effect of five N:P supply ratios and four growth rates (dilution rates). The relationship between cellular N and P quotas and growth rates fits well to both the Droop and Ågren’s functions for all species. We observed excess uptake of both N and P in the three species. N:P biomass ratios showed a significant positive relationship with N:P supply ratios across the entire range of growth rates, and N:P biomass ratios converged to an intermediate value independent of N:P supply ratios at higher growth rates. The effect of growth rates on N:P biomass ratios was positive at lower N:P supply ratios, but negative at higher N:P supply ratios for both Rhodomonas sp. and I. galbana, while for P. tricornutum this effect was negative at all N:P supply ratios. A significant interactive effect of N:P supply ratios and growth rates on N:P biomass ratios was found in both Rhodomonas sp. and P. tricornutum, but not in I. galbana. Our results suggest that Ågren’s functions may explain the underlying biochemical principle for the Droop model. The parameters in the Droop and Ågren’s functions can be useful indications of algal succession in the phytoplankton community in changing oceans.     

Optimization of growth conditions and fatty acid analysis for three freshwater diatom isolates

Diatoms are a group of highly abundant and diverse aquatic algae species. They contain high lipid content along with many bioactive compounds that can be exploited for biotechnological applications. Despite these attractive attributes, diatoms are underrepresented in production projects due to difficulties in their cultivation. To optimize the growth of three freshwater diatom isolates, Cyclotella sp., Synedra sp. and Navicula sp., an orthogonal assay on N, P, Si and Fe, as well as temperature and pH, was designed using traditional single‐factor tests. We also studied the effect of using nanosilica as an alternate Si source on growth and found that the diatom isolates studied achieved their highest growth rates under different combinations of nutrient and environmental conditions. Silica had the greatest influence on growth, followed by phosphate and iron. The optimized growth conditions for Synedra sp. were N: 30 mg L^?1, P: 3 mg L^?1, Si: 14.8 mg L^?1, Fe: 0.448 mg L^?1, temperature 25°C and pH 8. For Navicula sp.: N: 20 mg L^?1, P: 2.5 mg L^?1, Si: 19.7 mg L^?1, Fe: 0.112 mg L^?1, temperature 30°C and pH 7.5–8. For Cyclotella sp.: N: 20 mg L^?1, P: 2.5 mg L^?1, Si: 19.7 mg L^?1, Fe: 0.448 mg L^?1, temperature 30°C and pH 7.5–8. Nano silica negatively affected growth in Navicula sp. and Cyclotella sp., but no such effect was observed in Synedra sp. Fatty acid profiling showed C16:0, C16:1(n ? 7), C18:0 and C20:5(n ? 3) as major fatty acids, with no significant differences in fatty acid methyl ester profiles between traditional and modified media. This work gives us a new insight into the growth requirements of freshwater diatom species, which are less studied than marine species.     

Silicon application induces changes C:N:P stoichiometry and enhances stoichiometric homeostasis of sorghum and sunflower plants under salt stress

Beneficial effects of silicon (Si) on growth have been observed in some plant species, reportedly due to stoichiometric changes of C, N, and P. However, little is known about the effects on the stoichiometric relationships between C, N, and P when silicon is supplied via different modes in sorghum and sunflower plants under salt stress conditions. Therefore, the current study was performed to investigate the impact of differing modes of Si supply on shoot biomass production and C:N:P stoichiometry in sorghum and sunflower plants under salt stress. Two experiments were performed in a glass greenhouse using the strong Si-accumulator plant sorghum, as well as the intermediate type Si-accumulator sunflower, both of which were grown in pots filled with washed sand. Plant species were cultivated for 30 days in the absence or presence of salt stress (0 or 100 mM) and supplemented with one of four Si treatments: control plants (without Si), 28.6 mmol Si L⁻¹ via foliar application, 2.0 mmol Si L⁻¹ via nutrient solution, and combined application of foliar and nutrient solution, each group with five replications. The results revealed that supplied Si modified the C, N, and P concentrations, thereby enhancing the C:N:P stoichiometry and shoot dry matter of sorghum and sunflower plants under salt stress. Both application of Si via nutrient solution, as well as combined application via foliar and nutrient solution, increased the C:N ratio in both plant species under salt stress, but in sorghum plants decreased the C:P and N:P ratios and increased the shoot biomass production by 39%, while in sunflower plants increased the C:P and N:P ratios and increased the shoot biomass production by 24%. Our findings suggest that salt stress alleviation by Si impacts C:N:P stoichiometric relationships in a variable manner depending on the ability of the species to accumulate Si, as well as the route of Si administration.     

INTERACTIONS OF LIGHT/DARK CYCLES AND NITROGEN PULSES ON THE TIMING OF CELL DIVISION IN THE NITROGEN-LIMITED MARINE DIATOM CYLINDROTHECA FUSIFORMIS (BACILLARIOPHYCEAE)1

Cell division in most eukaryotic algae grown on alternating periods of light and dark (LD) is synchronized or phased so that cell division occurs only during a restricted portion of the LD cycle. However, the phase angle of the cell division gate, the time of division relative to the beginning of the light period, is known to be affected by growth conditions such as nutrient status and temperature. In this study, it is shown that the phase angle of cell division in a diatom, Cylindrotheca fusiformis Reimann and Lewin, is affected by the N-limited growth rate; cell division occurred later in the dark period (12:12 h LD cycle) when the growth rate was infradian (D = 0.42 d^?1) than when it was ultradian (D = 1.0 d^?1). Nitrogen-pulses did not affect the phase angle of the division gate, but could shift the time of peak cell division activity within the division gate. The effects, if any, of N-pulses were dependent upon the growth rate and the time of day that the pulses were administered. These responses indicate that the timing of cell division in this diatom is not determined solely by the zeitgeber from the LD cycle, but rather that a LD cycle control mechanism and a N-mediated control mechanism are both involved and are somewhat interdependent. In addition, an increase in protein was observed immediately after administering a N-pulse to C. fusiformis in the ultradian growth mode indicating that the accumulation of protein can be uncoupled from the cell division cycle.     

EFFECTS OF CADMIUM TOXICITY ON GROWTH AND ELEMENTAL COMPOSITION OF MARINE PHYTOPLANKTON

Some classes of marine phytoplankton are believed to be more tolerant of high concentrations of trace metals than others, but the results of experimental tests of this hypothesis are ambiguous. Eleven species of phytoplankton representing five classes were grown in Aquil medium containing Cd concentrations between 10⁻⁸ and 10⁻⁵ M ([Cd²⁺]= 10^−9.85 to 10^−6.84 M), and growth rates and intracellular concentrations of Cd, C, N, and S were measured. The mean Cd²⁺ concentration (pCd⁵⁰) that reduced the growth rate of each species to 50% of its maximum varied by 2.5 orders of magnitude, from 10^−6.23 for Emiliania huxleyi to 10^−8.79 for Synechococcus sp. Taxonomic trends in Cd resistance were not apparent in these data. Cadmium quotas (mol Cd·L⁻¹ cell volume) were lowest in species of Bacillariophyceae (ANOVA, P < 0.001), suggesting that they might regulate Cd transport differently than other taxa. Cellular S:C molar ratios increased in four of seven phytoplankton grown at high pCd (7.37–6.84) compared to low Cd ion concentrations (no added Cd), a result of increases in S·L⁻¹ cell volume. Nitrogen:carbon molar ratios were also higher in Cd-exposed phytoplankton, as changes in N and S were highly correlated ( r = 0.98, P < 0.0001). In two species that were examined, S:C ratios increased as a linear function of increasing Cd concentration. The results demonstrate large variability in Cd resistance among phytoplankton that is primarily a function of interspecific differences in Cd detoxification.     

NANOSTRUCTURE OF THE DIATOM FRUSTULE AS REVEALED BY ATOMIC FORCE AND SCANNING ELECTRON MICROSCOPY

The cell wall (frustule) of the freshwater diatom Pinnularia viridis (Nitzsch) Ehrenberg is composed of an assembly of highly silicified components and associated organic layers. We used atomic force microscopy (AFM) to investigate the nanostructure and relationship between the outermost surface organics and the siliceous frustule components of live diatoms under natural hydrated conditions. Contact mode AFM imaging revealed that the walls were coated in a thick mucilaginous material that was interrupted only in the vicinity of the raphe fissure. Analysis of this mucilage by force mode AFM demonstrated it to be a nonadhesive, soft, and compressible material. Application of greater force to the sample during repeated scanning enabled the mucilage to be swept from the hard underlying siliceous components and piled into columns on either side of the scan area by the scanning action of the tip. The mucilage columns remained intact for several hours without dissolving or settling back onto the cleaned valve surface, thereby revealing a cohesiveness that suggested a degree of cross-linking. The hard silicified surfaces of the diatom frustule appeared to be relatively smooth when living cells were imaged by AFM or when field-emission SEM was used to image chemically cleaned walls. AFM analysis of P. viridis frustules cleaved in cross-section revealed the nanostructure of the valve silica to be composed of a conglomerate of packed silica spheres that were 44.8 ± 0.7 nm in diameter. The silica spheres that comprised the girdle band biosilica were 40.3 ± 0.8 nm in diameter. Analysis of another heavily silicified diatom, Hantzschia amphioxys (Ehrenberg) Grunow, showed that the valve biosilica was composed of packed silica spheres that were 37.1 ± 1.4 nm and that silica particles from the girdle bands were 38.1 ± 0.5 nm. These results showed little variation in the size range of the silica particles within a particular frustule component (valve or girdle band), but there may be differences in particle size between these components within a diatom frustule and significant differences are found between species.     

Media optimization and lipid formation of two native diatoms for cultivation in the Southwest Texas desert

Two native diatom isolates belonging to the genus Nitzschia, TAMU-LBK-020 and TAMU-LBK-023, were grown in a chamber simulating summer temperatures and light:dark cycle in the southwest Texas (USA) desert. Different concentrations of macronutrients (Si, N, and P) were tested to identify conditions conducive to growth. Growth rates of the fastest growing treatments exceeded 0.6 doublings per day, with DW and AFDW exceeding 1.0 and 0.8 g L^?1, respectively. Treatments with positive growth were continued to a lipid formation step by omission of Si from the medium in this phase, with lipid accumulation monitored daily by Nile red fluorescence. Peak oil was observed at approximately day 3 for TAMU-LBK-020 and day 5 for TAMU-LBK-023. The highest oil concentrations observed were 17.5?±?7.3 % of AFDW in TAMU-LBK-020 and 28.5?±?5.6 % of AFDW in TAMU-LBK-023 at low N concentrations, but the low N also resulted in growth of 0.2 to 0.5 doublings per day. We propose a biphasic growth strategy, initially under a high macronutrient concentration to promote cell growth and division followed by starving for Si and N to achieve maximal lipid production. However, we anticipate that it will be necessary to determine optimal nutrient concentrations empirically for each species and strain. These results demonstrate that diatoms used for biofuels can achieve fast growth rates and high oil production, and also that native strains, when properly cultured, can be highly suitable for biofuel production, even in harsh climates such as the desert Southwest U.S.