Nitrate and ammonium absorption by plants growing at a sufficient or insufficient level of phosphorus in nutrient solutions

Summary Absorption of nitrate and ammonium was studied in water culture experiments with 4 to 6 weeks old plants of barley (Hordeum vulgare L.), buckwheat (Fagopyrum esculentum L. Moench) and rape (Brassica napus L.). The plants were grown in a complete nutrient solution with nitrate (5.7±0.2 mM) or nitrate (5.6±0.2 mM) + ammonium (0.04±0.02 mM). The pH of the nutrient solution was kept at 5.0 using a pH-stat. It was found that phosphorus deficiency reduced the rate of nitrate uptake by 58±3% when nitrate was the sole N source and by 83±1% when both nitrate and ammonium were present. The reduction occurred even before growth was significantly impeded by P deficiency. The inhibition of the uptake of ammonium was less,i.e. ammonium constituted 10±1% of the total N uptake in the P sufficient plants and 30±5% in the P deficient plants. The reduction of nitrate absorption greatly decreased the difference between the uptake of anions and cations. It is suggested that P deficiency reduced the assimilation of NO ₃ ^– into the proteins, which might cause a negative feedback on NO ₃ ^– influx and/or stimulate NO ₃ ^– efflux.     

Biofiltering efficiency in removal of dissolved nutrients by three species of estuarine macroalgae cultivated with sea bass (Dicentrarchus labrax) waste waters 2. Ammonium

Three estuarine macroalgae (Ulva rotundata,Enteromorpha intestinalis, Gracilariagracilis) of economic potential were cultivated in the laboratory toassess their biofiltering capacities for ammonium in waste effluents from a seabass (Dicentrarchus labrax) cultivation tank. The studywasdeveloped to investigate the functioning of N nutrition of the three species.Atlow water flow (< 2 volumes d^–1) the three species strippedefficiently the ammonium dissolved in the waste water from the fish tank, withaminimum biofiltering efficiency estimate of 61% in unstarved cultures ofG. gracilis at a water flow of 2 volumesd^–1. Maximum velocity for ammonium uptake (89.0 molNH₄ ⁺ g^–1 dry wth^–1) was found in U. rotundata,whereas G. gracilis showed the highest affinity for thisnutrient. The net ammonium uptake rate was significantly affected by the waterflow, being greatest at the highest flow assayed (2 volumesd^–1). Variations of tissue N and C:N ratios during aflow-through experiment suggested that N was not limiting macroalgal growth.However, when ammonium was supplied at a flow rate of 0.5 volumesd^–1, specially in a three-stage design, the marked reductionintissue N and the biomass C:N:P ratios suggested a more general nutrientdeficiency. A significant correlation was found between growth rates and the Nbiomass gained in the cultures. The three-stage design under low water flow(0.5volumes d^–1) showed that the highest ammonium uptake rates (upto 80.9 mol NH₄ ⁺ g^–1 dry wtd^–1 in U. rotundata) were found inthe first stage, with decreasing rates in the following ones. As a result, lowincrements or even losses of total N biomass in these stages were found,suggesting that ammonium was excreted from the algae. We conclude that thesespecies present a potential ability to biofilter the ammonium dissolved inwastewater from a D. labrax cultivation tank, suggesting thatscaling up the biofiltration designs, future practises using these macroalgaemay be implemented in the local fish farms, resulting in both environmental andeconomical advantages.     

Modeling of the pyruvate production with<Emphasis Type="Italic"> Escherichia coli</Emphasis> in a fed-batch bioreactor

A family of 10 competing, unstructured models has been developed to model cell growth, substrate consumption, and product formation of the pyruvate producing strain Escherichia coli YYC202 ldhA::Kan strain used in fed-batch processes. The strain is completely blocked in its ability to convert pyruvate into acetyl-CoA or acetate (using glucose as the carbon source) resulting in an acetate auxotrophy during growth in glucose minimal medium. Parameter estimation was carried out using data from fed-batch fermentation performed at constant glucose feed rates of q_VG=10 mL h^–1. Acetate was fed according to the previously developed feeding strategy. While the model identification was realized by least-square fit, the model discrimination was based on the model selection criterion (MSC). The validation of model parameters was performed applying data from two different fed-batch experiments with glucose feed rate q_VG=20 and 30 mL h^–1, respectively. Consequently, the most suitable model was identified that reflected the pyruvate and biomass curves adequately by considering a pyruvate inhibited growth (Jerusalimsky approach) and pyruvate inhibited product formation (described by modified Luedeking–Piret/Levenspiel term).List of symbols c_A acetate concentration (g L^–1) - c_A,0 acetate concentration in the feed (g L^–1) - c_G glucose concentration (g L^–1) - c_G,0 glucose concentration in the feed (g L^–1) - c_P pyruvate concentration (g L^–1) - c_P,max critical pyruvate concentration above which reaction cannot proceed (g L^–1) - c_X biomass concentration (g L^–1) - K_I inhibition constant for pyruvate production (g L^–1) - K_I^A inhibition constant for biomass growth on acetate (g L^–1) - K_P saturation constant for pyruvate production (g L^–1) - K_P inhibition constant of Jerusalimsky (g L^–1) - K_S^A Monod growth constant for acetate (g L^–1) - K_S^G Monod growth constant for glucose (g L^–1) - m_A maintenance coefficient for growth on acetate (g g^–1 h^–1) - m_G maintenance coefficient for growth on glucose (g g^–1 h^–1) - n constant of extended Monod kinetics (Levenspiel) (–) - q_V volumetric flow rate (L h^–1) - q_VA volumetric flow rate of acetate (L h^–1) - q_VG volumetric flow rate of glucose (L h^–1) - r_A specific rate of acetate consumption (g g^–1 h^–1) - r_G specific rate of glucose consumption (g g^–1 h^–1) - r_P specific rate of pyruvate production (g g^–1 h^–1) - r_P,max maximum specific rate of pyruvate production (g g^–1 h^–1) - t time (h) - V reaction (broth) volume (L) - Y_P/G yield coefficient pyruvate from glucose (g g^–1) - Y_X/A yield coefficient biomass from acetate (g g^–1) - Y_X/A,max maximum yield coefficient biomass from acetate (g g^–1) - Y_X/G yield coefficient biomass from glucose (g g^–1) - Y_X/G,max maximum yield coefficient biomass from glucose (g g^–1) - growth associated product formation coefficient (g g^–1) - non-growth associated product formation coefficient (g g^–1 h^–1) - specific growth rate (h^–1) - _max maximum specific growth rate (h^–1)     

Field energetics of a large carnivorous lizard,Varanus rosenbergi

Summary Field metabolic rates (FMRs; CO₂ production) and water influx rates of Varanus rosenbergi were measured seasonally by means of doubly-labelled water (³H₂ ¹⁸O). Metabolic rates and water influexes were highest in summer (0.18 ml CO₂ g^–1 h^–1 and 17.9 ml H₂O kg^–1 day^–1) and lowest in winter (0.04 ml CO₂ g^–1 h^–1 and 6.5 ml H₂O kg^–1 day^–1). FMRs and water fluxes were intermediate in spring. It is estimated that a 1-kg V. rosenbergi would need to consume 4.7 kg of prey each year to remain in energy balance and that the energy commitment to egg production requires a 40% increase in the food acquisition of a gravid female in late spring and early summer. It is calculated that water influxes in summer are totally provided from food but in other seasons, particularly winter, pulmocutaneous water exchange and/or drinking are significant avenues of water intake.     

The Quinol:Fumarate Oxidoreductase from the Sulphate Reducing Bacterium Desulfovibrio gigas: Spectroscopic and Redox Studies

The membrane bound fumarate reductase (FRD) from the sulphate-reducer Desulfovibrio gigas was purified from cells grown on a fumarate/sulphate medium and extensively characterized. The FRD is isolated with three subunits of apparent molecular masses of 71, 31, and 22 kDa. The enzyme is capable of both fumarate reduction and succinate oxidation, exhibiting a higher specificity toward fumarate (K _m for fumarate is 0.02 and for succinate 2 mM) and a reduction rate 30 times faster than that for oxidation. Studies by Visible and EPR spectroscopies allowed the identification of two B-type haems and the three iron–sulphur clusters usually found in FRDs and succinate dehydrogenases: [2Fe-2S]^2+/1+ (S1), [4Fe-4S]^2+/1+ (S2), and [3Fe-4S]^1+/0 (S3). The apparent macroscopic reduction potentials for the metal centers, at pH 7.6, were determined by redox titrations: –45 and –175 mV for the two haems, and +20 and –140 mV for the S3 and S1 clusters, respectively. The reduction potentials of the haem groups are pH dependent, supporting the proposal that fumarate reduction is associated with formation of the membrane proton gradient. Furthermore, co-reconstitution in liposomes of D. gigas FRD, duroquinone, and D. gigas cytochrome bd shows that this system is capable of coupling succinate oxidation with oxygen reduction to water.     

Biomass production in mixotrophic culture of Euglena gracilis under acidic condition and its growth energetics

When Euglena gracilis was grown in the heterotrophic condition with glucose and (NH₄)₂SO₄ as the carbon and nitrogen source, a high cell yield (4.28–4.48 g l^–1) was obtained and the culture pH decreased to 1.6–2. The biomass production in the heterotrophic culture was compared to those in the autotrophic and mixotrophic cultures. Autotrophic growth was 4.7–6.3% of the heterotrophic one, whereas about 15–19% higher growth was obtained in the mixotrophic culture. Moreover, good production of chlorophyll (39.4 mg l^–1) and carotenoids (13.8 mg l^–1) were attained in the mixotrophic culture, giving the highest fermenter productivity with respect to biomass as well as chlorophyll and carotenoids. Through an energetic analysis in the mixotrophic culture, it was estimated about 25–28% of the total ATP requirement is formed in the photochemical reactions. This resulted in an improved biomass production in the mixotrophic culture of E. gracilis.     

Production of S-adenosyl- L-methionine by a mutant strain of Kluyveromyces lactis

S-Adenosyl-l-methionine (AdoMet) was produced by a mutant strain Kluyveromyces lactis AM-65 grown on whey. A full factorial design method of three factors – (NH₄)₂SO₄ (factor x ₁), corn steep liquor (factor x ₂) and l-methionine (factor x ₃) on three levels – was used to determine the optimal medium conditions for the production of AdoMet. A time course shake-flask experiment in optimal whey medium (x ₁=3.1 g l^–1, x ₂=12.7 g l^–1, x ₃=4.6 g l^–1) was also carried out and the results confirmed the results of the factorial design and subsequent quadratic modelling and optimization of AdoMet production which reached 90 mg g^–1 cell dry wt.     

The production of cyclopiazonic acid by Penicillium commune and cyclopiazonic acid and aflatoxins by Aspergillus flavus as affected by water activity and temperature on maize grains

The combined effects of water activity (a_w) and temperature on mycotoxin production by Penicilium commune (cyclopiazonic acid — CPA) and Aspergillus flavus (CPA and aflatoxins — AF) were studied on maize over a 14-day period using a statistical experimental design. Analysis of variance showed a highly significant interaction (P 0.001) between these factors and mycotoxin production. The minimum a_w/temperature for CPA production (2264 ng g^–1 P. commune, 709 ng g^–1 A. flavus) was 0.90 a_w/30 °C while greatest production (7678 ng g^–1 P. commune, 1876 ng g^–1 A. flavus) was produced at 0.98 a_w/20 °C. Least AF (411 ng g^–1) was produced at 0.90 a_w/20 °C and most (3096 ng g^–1) at 0.98 a_w/30 °C.     

Enzyme production in continuous cultivation by the thermophilic fungus, Thermomyces lanuginosus

The production of extracellular enzymes by the thermophilic fungus Thermomyces lanuginosus was studied in chemostat cultures at a dilution rate of 0.08 h^–1 in relation to variation in the ammonium concentration in the feed medium. Under steady state conditions, three growth regimes were recognised and the production of several extracellular enzymes from T. lanuginosus was recorded under different nutrient limitations ranging from nitrogen limitation to carbon/energy limitation. The range and the production of carbohydrate hydrolysing enzymes and lipase increased from Regime I (NH₄Cl 600 mg l^–1) to Regime III (NH₄CI 1200 mg l^–1), whereas production of protease was highest in Regime II (600 mg l^–1 < NH₄Cl <1200 mg l^–1).     

Artemisinin production in Artemisia annua hairy root cultures with improved growth by altering the nitrogen source in the medium

Murashige & Skoog medium was modified for enhancing artemisinin production in Artemisia annua hairy root cultures by altering the ratio of NO ₃ ^– /NH ₄ ⁺ and the total amount of initial nitrogen. Increasing ammonium to 60 mM decreased both growth and artemisinin accumulation in hairy root cultures. With NO ₃ ^– /NH ₄ ⁺ at 5:1 (w/w), the optimum concentration of total initial nitrogen for artemisinin production was 20 mM. After 24 days of cultivation with 16.7 mM nitrate and 3.3 mM ammonium, the maximum artemisinin production of hairy roots was about 14 mg l^–1, a 57% increase over that in the standard MS medium.     

Promotion of indole alkaloid production in Catharanthus roseus cell cultures by rare earth elements

Production of the indole alkaloids, ajmalicine or catharanthine, in cell suspension cultures of Catharanthus roseus was enhanced by cerium (CeO₂ and CeCl₃), yttrium (Y₂O₃) and neodymium (NdCl₃). The yield of ajmalicine in these treated-cultures reached 51 mg l^–1 (CeO₂), 40 mg l^–1 (CeCl₃), 41 mg l^–1 (Y₂O₃) and 49 mg l^–1 (NdCl₃) while catharanthine production reached to 36 mg l^–1 (CeO₂) and 31 mg l^–1 (CeCl₃). A major portion of increased alkaloids was released into medium in these treatments. But Sm₂O₃, SmCl₃, La₂O₃, LaCl₃, complex of chromium (III)-titanium (IV) and NaSeO₄ treatments had little effect on alkaloid production of C. roseus cell cultures.     

Modelling continuous culture of Rhodospirillum rubrum in photobioreactor under light limited conditions

Rhodospirillum rubrum was grown continuously and photoheterotrophically under light limitation using a cylindrical photobioreactor in which the steady state biomass concentration was varied between 0.4 to 4 kg m^–3 at a constant radiant incident flux of 100 W m^–2. Kinetic and stoichiometric models for the growth are proposed. The biomass productivities, acetate consumption rate and the CO₂ production rate can be quantitatively predicted to a high level of accuracy by the proposed model calculations. Nomenclature: C _X, biomass concentration (kg m^–3) D, dilution rate (h^–1) Ea, mean mass absorption coefficient (m² kg^–1) I , total available radiant light energy (W m^–2) K, half saturation constant for light (W m^–2) R _W, boundary radius defining the working illuminated volume (m) r _X, local biomass volumetric rate (kg m^–3 h^–1) <r _X>, mean volumetric growth rate (kg m^–3 h^–1) V _W, illuminated working volume in the PBR (m^–3). Greek letters: , working illuminated fraction (–) _M, maximum quantum yield (–) bar, mean energetic yield (kg J^–1).     

Levan production using mutant strains of Zymomonas mobilis in different culture conditions

Several levan hyperproducing mutants of Zymomonas mobilis strains were selected by mutagenesis with N-methyl-N-nitro-nitrosoguanidine and caffeine. Highest levan production (41 g l^–1) was obtained with a mutant strain HL 29 in a culture medium containing 200 g sucrose l^–1 and 0.5 g (NH₄)₂SO₄ l^–1 stored at 7 °C for 29 days. This is the first report describing the levan synthesis by Z. mobilis at 7 °C.     

Ulva rigida (Ulvales,Chlorophyta) tank culture as biofilters for dissolved inorganic nitrogen from fishpond effluents

Ulva rigida was cultivated in 7501 tanks at different densities with direct and continuous inflow (at 2, 4, 8 and 12 volumes d^–1) of the effluents from a commercial marine fishpond (40 metric tonnes, Tm, of Sparus aurata, water exchange rate of 16 m³ Tm^–1) in order to assess the maximum and optimum dissolved inorganic nitrogen (DIN) uptake rate and the annual stability of the Ulva tank biofiltering system. Maximum yields (40 g DW m^–2 d^–1) were obtained at a density of 2.5 g FW 1^–1 and at a DIN inflow rate of 1.7 g DIN m^–2 d^–1. Maximum DIN uptake rates were obtained during summer (2.2 g DIN M^–2 d^–1), and minimum in winter (1.1 g DIN m^–2 d^–1) with a yearly average DIN uptake rate of 1.77 g DIN m^–2 d^–1 At yearly average DIN removal efficiency (2.0 g DIN m^–2 d^–1, if winter period is excluded), 153 m² of Ulva tank surface would be needed to recover 100% of the DIN produced by 1 Tm of fish.Abbreviations DIN= dissolved inorganic nitrogen (NH _inf4 ^sup+ + NO _inf3 ^sup– + NO _inf2 ^sup– ); - FW= fresh weight; - DW= dry weight; - PFD= photon flux density; - V= DIN uptake rate     

Ammonium and nitrate uptake in gap,generalist and understory species of the genus Piper

Summary We studied root net uptake of ammonium (NH ₄ ⁺ ) and nitrate (NO ₃ ^– ) in species of the genus Piper (Piperaceae) under high, intermediate and low photosynthetically active photon flux densities (PFD). Plants were grown hydroponically, and then transferred to temperature controlled (25° C) root cuvettes for nutrient uptake determinations. Uptake solutions provided NH ₄ ⁺ and NO ₃ ^– simultaneously (both) or separately (single). In the first experiment, seven species of Piper, from a broad range of rainforest light habitats ranging from gap to understory, were screened for mineral nitrogen preference (100 M NH ₄ ⁺ and/or 100 M NO ₃ ^– ) at intermediate PFD (100 mol m^–2 s^–1). Preference for NH ₄ ⁺ relative to NO ₃ ^– , defined as the ratio of NH ₄ ⁺ (both):NO ₃ ^– (both) net uptake, was higher in understory species than in gap species. Ammonium repression of NO ₃ ^– uptake, defined as the ratio of NO ₃ ^– (single): NO ₃ ^– (both) net uptake, was also higher in understory species as compared to gap species. In a second set of experiments, we examined the effect of nitrogen concentration (equimolar, 10 to 1000 M) on NH ₄ ⁺ preference and NH ₄ ⁺ repression of NO ₃ ^– net uptake at high (500 mol m^–2 s^–1) and low (50 mol m^–2 s^–1) PFD in a gap (P. auritum), generalist (P. hispidum) and understory species (P. aequale). All species exhibited negligible NH ₄ ⁺ repression of NO ₃ ^– net uptake at high PFD. At low PFD, NH ₄ ⁺ preference and repression of NO ₃ ^– net uptake occurred in all species (understory > generalist > gap), but only at intermediate nitrogen concentrations, i.e. between 10 and 200 M. Ammonium repression of net NO ₃ ^– uptake decreased or increased rapidly (in < 48 h) after transitions from low to high or from high to low PFD respectively. No significant diurnal patterns in NO ₃ ^– or NH ₄ ⁺ net uptake were observed.CIWDPB publication # 1130     

Water treatment of land-based fish farm effluents by outdoor culture of marine diatoms

The feasibility of using fish farm effluents was evaluated as a source of inorganic nutrients for mass production of marine diatoms. Batch cultures were conducted from May to July 1995 in 16-L outdoor rectangular tanks, homogenized by gentle aeration (0.2 L _air L^–1 h^–1). The effluents from the two fish farms studied were both characterized by high concentrations of inorganic materials (NH₄-N, PO₄-;P, Si(OH)₄-Si) and were shown to support production of marine diatoms. Moreover, periodic measurements of inorganic matter levels in the cultures showed that clearance was efficacious (90% in 3–5 days). Water purification efficiency and culture productivity were further increased through appropriate nutrient balancing. When effluents were limited in silicate, addition of Na₂SiO₃ induced a significant increase in both diatom biomass and nutrient removal efficiency. In this case, up to 720 000 cell mL^–1 were produced dominated bySkeletonema costatum. By contrast, in effluents loaded with silicate, adjustment of the N:P:Si ratio by NH₄-N and PO₄-P supplementation then gave increased biomass production. In this case, the maximum cell density found was 450 000 cell mL^–1, dominated byChaetoceros spp.Author for correspondence     

Selective production of bikaverin in a fluidized bioreactor with immobilized Gibberella fujikuroi

The best culture medium composition for the production of bikaverin by Gibberella fujikuroi in shake-flasks, i.e. 100 g glucose l^–1; 1 g NH₄Cl l^–1; 2 g rice flour l^–1; 5 g KH₂PO₄ l^–1 and 2.5 g MgSO₄ l^–1, was obtained through a fractional factorial design and then scaled-up to a fluidized bioreactor. The effects of carbon and nitrogen concentrations, inoculum size, aeration, flow rate and bead sizes on batch bikaverin production using immobilized G. fujikuroi in a fluidized bioreactor were determined by an orthogonal experimental design. Concentrations of up to 6.83 g bikaverin l^–1 were obtained when the medium contained 100 g glucose l^–1 and 1 g NH₄Cl l^–1 with an inoculum ratio of 10% v/v, an aeration rate of 3 volumes of air per volume of medium min^–1, and a bead size of 3 mm. Based on dry weight, the bikaverin production was 30–100 times larger than found in submerged culture and approximately three times larger than reported for solid substrate fermentation.     

Biofiltering efficiency in removal of dissolved nutrients by three species of estuarine macroalgae cultivated with sea bass (Dicentrarchus labrax) waste waters 1. Phosphate

The potential of three estuarine macroalgae (Ulvarotundata, Enteromorpa intestinalis andGracilaria gracilis) as biofilters for phosphate ineffluents of a sea bass (Dicentrarchus labrax) cultivationtank was studied. These seaweeds thrive in Cádiz Bay and were alsoselected because of their economic potential, so that environmental andeconomicadvantages may be achieved by future integrated aquaculture practices in thelocal fish farms. The study was designed to investigate the functioning of Pnutrition of the selected species. Maximum velocity of phosphate uptake (2.86mol PO₄ g^–1 dry wth^–1) was found in U. rotundata.This species also showed the highest affinity for this nutrient. At low flowrates (< 2 volumes d^–1), the three species efficientlyfiltered the phosphate dissolved in the waste water, with a minimum efficiencyof 60.7% in U. rotundata. Net phosphate uptake rate wassignificantly affected by the water flow, being greatest at the highest rateassayed (2 volumes d^–1). The marked decrease in tissue P shownby the three species during a flow-through experiment suggested that growth wasP limited. However, due to the increase in biomass, total P biomass increasedinthe cultures. A significant correlation was found between growth rates and thenet P biomass gained in the cultures. A three-stage design under low water flow(0.5 volumes d^–1) showed that the highest growth rates (up to0.14 d^–1) and integrated phosphate uptake rates(up to 5.8 mol PO₄ ^3– g^–1dry wt d^–1) were found in E.intestinalis in the first stage, with decreasing rates in thefollowing ones. As a result, phosphate become limiting and low increments oreven losses of total P biomass in these stages were found suggesting thatphosphate was excreted from the algae. The results show the potential abilityofthe three species to reduce substantially, at low water flow, the phosphateconcentration in waste waters from a D. labrax cultivationtank, and thus the quality of effluents from intensive aquaculture practices.     

Interactive effects of exogenous combined nitrogen and phosphorus on growth and nitrogen fixation by azolla

Effects of N source and media-N and P levels were examined on growth, N uptake, and N₂ fixation ofAzolla pinnata withAnabaena azollae association (azolla) at two inoculum-P concentrations. Each expeiment was conducted for 7 days in a growth chamber using azolla at a predetermined inoculum-P concentration and the growth media containing a combination of four levels of P (0, 15, 75, and 200 M) and three levels (0, 1, and 5 mM) of either¹⁵N-enriched NH ₄ ⁺ as ammonium sulfate or¹⁵N-enriched NO ₃ ^– as potassium nitrate. Nitrogen uptake and N₂ fixation were measured by¹⁵N isotopic dilution method. Tissue P and N, N uptake, and N₂-fixation increased with increasing P concentration in the media regardless of the inoculum-P level of azolla. Increasing P concentration in the media increased growth of azolla at low inoculum P, but the effect on high inoculum-P azolla was either small or absen. High inoculum-P concentration resulted in increased growth, tissue-N and P concentrations, N uptake, and N₂ fixation by azolla. Ammonium in the growth media caused larger increase in tissue-N and greater repression of N₂ fixation than equimolar concentration of NO ₃ ^– . In the presence of NH ₄ ⁺ or NO ₃ ^– , in the growth media, N uptake by azolla exceeded the corresponding decrease in N₂ fixation, resulting in an overall increase in tissue-N concentration. Phosphorus in the media tended to negate the inhibitory effect of NH ₄ ⁺ or NO ₃ ^– on N₂ fixation. A multiple regression model showed that the effect of tissue-N on N₂ fixation was negative while that of tissue-P was positive. Therefore, a relative change in tissue-N and P appeared to regulate N₂ fixation. Tissue-N and P had similar effects on relative growth rate of azolla also. Inoculum-P level of azolla was important in determining the response to media-P.This research was supported by a grant from USAID under Indo-US Science and Technology Initiative.     

Optimization of cell growth and poly(3-hydroxybutyrate) accumulation on date syrup by a Bacillus megaterium strain

Optimal growth and PHB accumulation in Bacillus megaterium occurred with 5% (w/v) date syrup or beet molasses supplemented with NH₄Cl. When date syrup and beet molasses were used alone without an additional nitrogen source, a cell density of about 3gl^–1 with a PHB content of the cells of 50% (w/w) was achieved. NH₄NO₃ followed by ammonium acetate and then NH₄Cl supported cell growth up to 4.8gl^–1, whereas PHB accumulation was increased with NH₄Cl followed by ammonium acetate, NH₄NO₃ and then (NH₄)₂SO₄ to a PHB content of nearly 42% (w/w). Cultivation of B.megaterium at 30l scale gave a PHB content of 25% (w/w) of the cells and a cell density of 3.4gl^–1 after 14h growth.