Increased CO<Subscript>2</Subscript> and the effect of pH on growth and calcification of <Emphasis Type="Italic">Pleurochrysis carterae</Emphasis> and <Emphasis Type="Italic">Emiliania huxleyi</Emphasis> (Haptophyta) in semicontinuous cultures

The effects of changes in CO₂ and pH on biomass productivity and carbon uptake of Pleurochrysis carterae and Emiliania huxleyi in open raceway ponds and a plate photobioreactor were studied. The pH of P. carterae cultures increased during day and decreased at night, whereas the pH of E. huxleyi cultures showed no significant diurnal changes. P. carterae coccolith production occurs during the dark period, whereas in E. huxleyi, coccolith production is mainly during the day. Addition of CO₂ at constant pH (pH-stat) resulted in an increase in P. carterae biomass and coccolith productivity, while CO₂ addition lowered E. huxleyi biomass and coccolith production. Neither of these algae could grow at less than pH 7.5. Species-specific diurnal pH and pCO₂ variations could be indicative of significant differences in carbon uptake between these two species. While E. huxleyi has been suggested to be predominantly a bicarbonate user, our results indicate that P. carterae may be using CO₂ as the main C source for photosynthesis and calcification.     

Hydroxylation of naphthalene by aromatic peroxygenase from <Emphasis Type="Italic">Agrocybe aegerita</Emphasis> proceeds via oxygen transfer from H<Subscript>2</Subscript>O<Subscript>2</Subscript> and intermediary epoxidation

Agrocybe aegerita peroxidase/peroxygenase (AaP) is an extracellular fungal biocatalyst that selectively hydroxylates the aromatic ring of naphthalene. Under alkaline conditions, the reaction proceeds via the formation of an intermediary product with a molecular mass of 144 and a characteristic UV absorption spectrum (A _max 210, 267, and 303 nm). The compound was semistable at pH 9 but spontaneously hydrolyzed under acidic conditions (pH <7) into 1-naphthol as major product and traces of 2-naphthol. Based on these findings and literature data, we propose naphthalene 1,2-oxide as the primary product of AaP-catalyzed oxygenation of naphthalene. Using ¹⁸O-labeled hydrogen peroxide, the origin of the oxygen atom transferred to naphthalene was proved to be the peroxide that acts both as oxidant (primary electron acceptor) and oxygen source.     

Productivity and biochemical composition of <Emphasis Type="Italic">Tetradesmus obliquus</Emphasis> and <Emphasis Type="Italic">Phaeodactylum tricornutum</Emphasis>: effects of different cultivation approaches

The present work evaluated biomass productivity, carbon dioxide fixation rate, and biochemical composition of two microalgal species, Phaeodactylum tricornutum (Bacillariophyta) and Tetradesmus obliquus (Chlorophyta), cultivated indoors in high-technology photobioreactors (HT-PBR) and outdoors both in pilot ponds and low-technology photobioreactors in a greenhouse in southern Italy. Microalgae were grown in standard media, under nitrogen starvation, and in two liquid digestates obtained from anaerobic digestion of agro-zootechnical and vegetable biomass. P. tricornutum, cultivated in semi-continuous mode in indoor HT-PBRs with standard medium, showed a biomass productivity of 21.0?±?2.3 g m^?2 d^?1. Applying nitrogen starvation, the lipid productivity increased from 2.3 up to 4.5?±?0.5 g m^?2 d^?1, with a 24 % decrease of biomass productivity. For T. obliquus, a biomass productivity of 9.1?±?0.9 g m^?2 d^?1 in indoor HT-PBR was obtained using standard medium. Applying liquid digestates as fertilizers in open ponds, T. obliquus gave a biomass productivity (10.8?±?2.0 g m^?2 d^?1) not statistically different from complete medium such as P. tricornutum (6.5?±?2.2 g m^?2 d^?1). The biochemical data showed that the fatty acid composition of the microalgal biomass was affected by the different cultivation conditions for both microalgae. In conclusion, it was found that the microalgal productivity in standard medium was about doubled in HT-PBR compared to open ponds for P. tricornutum and was about 20 % higher for T. obliquus.     

Influence of flow turbulence on growth and indole acetic acid and H<Subscript>2</Subscript>O<Subscript>2</Subscript> metabolism of three aquatic macrophyte species

This study aimed to determine the influence of different turbulence levels on morphologically different aquatic macrophyte species. The variations of growth and the alterations of indole acetic acid (IAA) and H₂O₂ metabolism of three morphologically distinct aquatic macrophyte species, Elodea nuttallii, Potamogeton crispus and Vallisneria spiralis were studied in the exposure to three different turbulence conditions. Plants were subjected to turbulence generated by vertically oscillating horizontal grids at three different frequencies: 1, 2 and 4 Hz. A no-turbulence condition was used as a control. The growth of V. spiralis and E. nuttallii was suppressed by exposure to greater turbulence, though increased turbulence enhanced the growth of P. crispus. The growth rate of plants was positively correlated with IAA concentration. IAA concentration was negatively correlated with turbulence level in V. spiralis and E. nuttallii, while it was positively correlated in P. crispus. H₂O₂ concentration was positively correlated with turbulence level in the former species, while it had no correlation in the latter. P. crispus showed more resistance to turbulence-induced mechanical stress and growth suppression than the other two species. The observed variations of growth and IAA and H₂O₂ metabolism showed that plant morphology has a significant influence on the response to flow turbulence. The results from this study provide a basis for understanding the mechanism and consequential morphological changes in plants that grow in moving water.     

Toxicity of cadmium and zinc on two microalgae, <Emphasis Type="Italic">Scenedesmus obliquus</Emphasis> and <Emphasis Type="Italic">Desmodesmus pleiomorphus</Emphasis>, from Northern Portugal

Aquatic environments often contain toxic heavy metals that may enter the food web via uptake by microalgae and eventually cause severe poisoning problems at higher trophic levels. The effects of Cd and Zn cations upon growth of two native green microalgal species, Scenedesmus obliquus and Desmodesmus pleiomorphus (previously isolated from a polluted site in Northern Portugal), were accordingly evaluated. Growth inhibition of the microalgal cells was determined following exposure for 96 h to several initial concentrations of aqueous solutions of either of those two metals. At the higher end of Cd and Zn experimental concentration ranges, a significant reduction in cell density was observed in the cultures; EC₅₀ values, calculated after fitting a Weibull model to the experimental data, were 0.058 and 1.92 mg L⁻¹ for Cd and 16.99 and 4.87 mg L⁻¹ for Zn in the case of S. obliquus and D. pleiomorphus, respectively. One observed that S. obliquus can tolerate higher Zn concentrations than D. pleiomorphus, but the reverse holds regarding exposure to Cd.     

Skeletal alterations and polymorphism in a Mediterranean bryozoan at natural CO<Subscript>2</Subscript> vents

Colonies of the cheilostome bryozoan Schizoporella errata were grown at a site near Ischia Island (Tyrrhenian Sea, Italy) where volcanogenic CO₂ emissions lower seawater pH to 7.76, simulating levels of ocean acidification predicted for the end of the present century. Compared with colonies from a control site (mean pH = 8.09), putative defensive polymorphs (avicularia) were significantly fewer, and retarded growth of zooidal basal and lateral walls was evident at the low pH site. The lower proportion of avicularia suggests a switch in resource allocation away from defence to favouring rapid growth. In addition, corrosion of the skeleton was observed in both new and old zooids at the low pH site, and feeding zooids were slightly smaller but had larger orifices for the protrusion of feeding lophophores. These findings corroborate previous studies demonstrating potential dissolution of carbonate skeletons in low pH seawater, while providing new insight into the possible ability of colonial species to respond to ocean acidification by adjusting resource allocation between zooids of different types.     

The physiological response of marine diatoms to ocean acidification: differential roles of seawater pCO2 and pH

Although increasing the pCO₂ for diatoms will presumably down‐regulate the CO₂‐concentrating mechanism (CCM) to save energy for growth, different species have been reported to respond differently to ocean acidification (OA). To better understand their growth responses to OA, we acclimated the diatoms Thalassiosira pseudonana, Phaeodactylum tricornutum, and Chaetoceros muelleri to ambient (pCO₂ 400 μatm, pH 8.1), carbonated (pCO₂ 800 μatm, pH 8.1), acidified (pCO₂ 400 μatm, pH 7.8), and OA (pCO₂ 800 μatm, pH 7.8) conditions and investigated how seawater pCO₂ and pH affect their CCMs, photosynthesis, and respiration both individually and jointly. In all three diatoms, carbonation down‐regulated the CCMs, while acidification increased both the photosynthetic carbon fixation rate and the fraction of CO₂ as the inorganic carbon source. The positive OA effect on photosynthetic carbon fixation was more pronounced in C. muelleri, which had a relatively lower photosynthetic affinity for CO₂, than in either T. pseudonana or P. tricornutum. In response to OA, T. pseudonana increased respiration for active disposal of H⁺ to maintain its intracellular pH, whereas P. tricornutum and C. muelleri retained their respiration rate but lowered the intracellular pH to maintain the cross‐membrane electrochemical gradient for H⁺ efflux. As the net result of changes in photosynthesis and respiration, growth enhancement to OA of the three diatoms followed the order of C. muelleri > P. tricornutum > T. pseudonana. This study demonstrates that elucidating the separate and joint impacts of increased pCO₂ and decreased pH aids the mechanistic understanding of OA effects on diatoms in the future, acidified oceans.     

Identifying non-invasible habitats for marine copepods using temperature-dependent <Emphasis Type="Italic">R</Emphasis><Subscript>0</Subscript>

If a non-indigenous species is to thrive and become invasive it must first persist under its new set of environmental conditions. Net reproductive rate (R ₀) represents the average number of female offspring produced by a female over its lifetime, and has been used as a metric of population persistence. We modeled R ₀ as a function of ambient water temperature (T) for the invasive marine calanoid copepod Pseudodiaptomus marinus, which is introduced to west coast of North America from East Asia by ship ballast water. The model was based on temperature-dependent stage-structured population dynamics given by a system of ordinary differential equations. We proposed a methodology to identify habitats that are non-invasible for P. marinus using the threshold of R ₀(T) < 1 in order to identify potentially invasible habitats. We parameterized the model using published data on P. marinus and applied R ₀(T) to identify the range of non-invasible habitats in a global scale based on sea surface temperature data. The model predictions matched the field evidence of species occurrences well.     

Efficient separation and purification of allophycocyanin from <Emphasis Type="Italic">Spirulina</Emphasis> (<Emphasis Type="Italic">Arthrospira</Emphasis>) <Emphasis Type="Italic">platensis</Emphasis>

Allophycocyanin (APC) is a minor component of phycobiliproteins in cyanobacteria and red algae. This paper describes a simple and inexpensive extracting method for isolating APC from Spirulina (Arthrospira) platensis with high efficiency. The crude phycobiliprotein extract was pretreated by ammonium sulfate fractionation. Then, by adding hydroxylapatite into crude phycobiliprotein extract dissolved in 20 mM phosphate buffer (pH 7.0), APC was selectively adsorbed by hydroxylapatite but C-phycocyanin (C-PC) was not. The hydroxylapatite was collected and APC was extracted from the crude phycobiliprotein extract. Then, the enriched APC was washed off from the hydroxylapatite using 100 mM phosphate buffer (pH 7.0). In this simple extracting method it was easy to remove C-PC and isolate APC in large amounts. The absorbance ratio A ₆₅₀/A ₂₈₀ of extracted APC reached 2.0. The recovery yield was 70%, representing 4.61 mg · g⁻¹ wet weight. The extracted APC could be further purified by a simple anion-exchange chromatography with a pH gradient from 5.6 to 4.0. The absorbance ratio A ₆₅₀/A ₂₈₀ of the purified APC reached 5.0, and the overall recovery yield was 43%, representing 2.83 mg · g⁻¹ wet weight. Its purity was confirmed by native polyacrylamide gel electrophoresis (PAGE) and sodium dodecyl sulfate-PAGE.     

Magnetophoretic harvesting of freshwater microalgae using polypyrrole/Fe<Subscript>3</Subscript>O<Subscript>4</Subscript> nanocomposite and its reusability

A magnetophoretic harvesting agent, a polypyrrole/Fe₃O₄ magnetic nanocomposite, is proposed as a cost and energy efficient alternative to recover biomass of the microalgae Botryococcus braunii, Chlorella protothecoides, and Chlorella vulgaris from their culture media. The maximal recovery efficiency reached almost 99 % for B. braunii, 92.4 % for C. protothecoides, and 90.8 % for C. vulgaris. The maximum adsorption capacity (Q ₀) of the magnetic nanocomposite for B. braunii (63.49 mg dry biomass mg^?1 PPy/Fe₃O₄) was higher than that for C. protothecoides (43.91 mg dry biomass mg^?1 PPy/Fe₃O₄) and C. vulgaris (39.98 mg dry biomass mg^?1 PPy/Fe₃O₄). The highest harvesting efficiency for all the studied microalgae were at pH 10.0, and measurement of zeta-potential confirmed that the flocculation was induced by charge neutralization. This study showed that polypyrrole/Fe₃O₄ can be a promising flocculant due to its high efficacy, low dose requirements, short settling time, its integrity with cells, and with great potential for saving energy because of its recyclability.     

Evaluation of electrocoagulation,flocculation, and sedimentation harvesting methods on microalgae consortium grown in anaerobically digested abattoir effluent

Microalgae dewatering is a major bottleneck for biomass production in a large-scale microalgal production system which accounts for 20–60% of production cost. In this study, three dewatering systems of electrocoagulation, flocculation, and pH-induced flocculation were evaluated for microalgal consortium grown in anaerobically digested abattoir effluent at pH 6.5 and 9.5. At the shortest time (15 min) and the highest current density (0.08 A cm^?2), the highest microalgae recoveries of 78 and 84% were obtained with the corresponding power consumptions of 1.25 and 1.07 kWh kg^?1 for cultures at pH 6.5 and 9.5. For microalgae suspension at pH 6.5, the highest biomass recovery of 77% was obtained when 100 mg L^?1 of FeCl₃·6H₂O (after 15 min) or 100 mg L^?1 of Al₂(SO₄)₃·18H₂O (after 30 min) was added. However, microalgal recoveries significantly increased when FeCl₃·6H₂O or Al₂(SO₄)₃·18H₂O was used with the culture at pH 9.5. pH-Induced experiments showed that cultures adjusted at pH 10.5 had 36% higher biomass recovery compared to that in cultures at pH 8.5 after 2 h. The results of this study showed that cultures at higher pH (9.5) had a better microalgae recovery in all dewatering systems than cultures at lower pH (6.5).

     

Effects of pH and bicarbonate on the nutrient status and growth of three Lupinus species

Aims

High pH, and high bicarbonate (HCO₃⁻) and calcium (Ca) availability characterise calcareous soils. High [Ca] only partially explains why some Lupinus species are calcifuge, so we explored high [HCO₃⁻] and high pH.

Methods

We grew six Lupinus genotypes in hydroponics with pH 5, 6.5 and 8^a (adjusted by KOH), and 8^b (adjusted by KHCO₃). Leaf symptoms and areas, root appearance and biomass were recorded; whole leaf and root nutrient concentrations, and leaf cellular phosphorus (P), Ca and potassium (K) concentrations were determined using elemental X-ray microanalysis.

Results

Chlorosis was observed in young leaves at high pH for L. angustifolius and L. cosentinii, and P deficiency at high pH for all genotypes. High pH decreased iron (Fe) and zinc (Zn) uptake in all genotypes. It also decreased lateral root growth, the uptake of P, K, Ca, and manganese (Mn) by all sensitive species; and translocation of P, Fe, Zn, Mn, and Ca to leaves in most sensitive species. However, leaf [Ca], leaf [K], [K] within each measured cell type, and translocation of K and Ca to leaves of L. pilosus and L. cosentinii at pH 8 were greater than at pH 5 and 6.5. Compared with pH 8^a, all L. angustifolius genotypes translocated more P, Fe, Zn, Mn and K from roots to leaves at pH 8^b. High pH did not affect the leaf cell types that accumulated P and Ca, but decreased the leaf cellular [P].

Conclusions

Lupinus angustifolius and L. cosentinii were sensitive to high [HCO₃⁻] and/or high pH; L. pilosus was relatively tolerant. High pH decreased lateral root growth and nutrient uptake, inhibiting growth of sensitive species. High [HCO₃⁻] diminished the negative effect of pH 8 on nutrient translocation to leaves in most L. angustifolius genotypes. This knowledge provides critical insights into the habits of Lupinus species to guide breeding of calcicole plants.

     

Effective biomass harvesting of marine diatom Chaetoceros muelleri by chitosan-induced flocculation,preservation of biomass,and recycling of culture medium for aquaculture feed application

Microalgae are a promising new source of biomass; however, large-scale economical harvesting of microalgal biomass is a major technological and economic challenge, limiting the commercial production of microalgal biomass for high-value compounds. In this study, the cationic polymer chitosan was used for the harvesting of the marine diatom Chaetoceros muelleri. Natural flocculation, and pH and chitosan-induced flocculation were studied in detail. The effects of flocculant dosage, culture pH, initial biomass concentration, and sedimentation time were investigated on biomass recovery. The results showed that flocculation efficiency can reach > 99% with an optimum dosage of chitosan (80 mg L^?1) at pH 9.6 and settling time of 40 minutes for biomass concentration from 0.2 to 1.2 g L^?1. The reusability of the recycled water, preservation of biomass after harvesting, and cost of the harvesting process were evaluated. The results showed that the chitosan-induced flocculation offers an efficient, cost-effective, rapid, and sustainable harvesting method for C. muelleri biomass for food and feed applications in aquaculture.

     

High-level heterologous expression of an alkaline lipase gene from <Emphasis Type="Italic">Penicillium cyclopium</Emphasis> PG37 in <Emphasis Type="Italic">Pichia pastoris</Emphasis>

A 777-bp cDNA fragment encoding a mature alkaline lipase (LipI) from Penicillium cyclopium PG37 was amplified by RT–PCR, and inserted into the expression plasmid pPIC9 K. The recombinant plasmid, designated as pPIC9 K-lipI, was linearized with SalI and transformed into Pichia pastoris GS115 (his4, Mut⁺) by electroporation. MD plate and YPD plates containing G418 were used for screening of the multi-copy P. pastoris transformants (His⁺, Mut⁺). One transformant resistant to 4.0 mg/ml of G418, numbered as P. pastoris GSL4-7, expressing the highest recombinant LipI (rLipI) activity was chosen for optimizing expression conditions. The integration of the gene LipI into the P. pastoris GS115 genome was confirmed by PCR analysis using 5′- and 3′-AOX1 primers. SDS–PAGE and lipase activity assays demonstrated that the rLipI, a glycosylated protein with an apparent molecular weight of about 31.5 kDa, was extracellularly expressed in P. pastoris. When the P. pastoris GSL4-7 was cultured under the optimized conditions, the expressed rLipI activity was up to 407 U/ml, much higher than that (10.5 U/ml) expressed with standard protocol. The rLipI showed the highest activity at pH 10.5 and 25°C, and was stable at a broad pH range of 7.0–10.5 and at a temperature of 30°C or below.     

Changes in variable charge and acidity of rye (<Emphasis Type="Italic">Secale cereale</Emphasis> L<Emphasis Type="Italic">.</Emphasis>) roots surface under Zn-stress

The plants of winter rye variety Rostockie grown in nutrient solution prepared according to Marschner and Romheld (1983) were stressed at shooting stage with different zinc (ZnCl₂) concentrations of 0, 20, 200 and 400 mgXdm⁻³ during 10 days at pH = 4.5. The control plants were grown continuously at pH = 7, without Zn²⁺. The roots of all plants were titrated with 0.1 molXdm⁻³ NaOH in 1 molXdm⁻³ NaCl solution with a rate 0.01 ml/min using auto-titrator Titrino 702 (Metrohm). The amount of the base consumed between pH 3 and 10 was recorded with a step of 0.1 pH unit. From the titration data root surface charge was characterized. Roots grown at pH = 4.5 without Zn²⁺ addition, had apparently the same charge properties as the control roots. Under 200 and 400 mgXdm⁻³ of zinc addition, variable charge, Q, and the cation exchange capacity, CEC, of the roots significantly decreased as did the fractions of surface acidic functional groups of strong and medium acidity i.e. the groups having apparent surface dissociation constants, Kapp, between 10^−3.5 and 10^−5.5. For 20 mgXdm⁻³ zinc stress the fraction of weaker surface groups (surface dissociation constants around 10^−6.5) decreased also. The surface charge density, SCD, of roots decreased under the Zn stress.     

The Effects of Copper (II) Ions on <Emphasis Type="Italic">Enterococcus hirae</Emphasis> Cell Growth and the Proton-Translocating F<Subscript>o</Subscript>F<Subscript>1</Subscript> ATPase Activity

Enterococcus hirae grow well under anaerobic conditions at alkaline pH (pH 8.0) producing acids by glucose fermentation. Bacterial growth was shown to be accompanied by decrease of redox potential from positive values (~+35 mV) to negative ones (~−220 mV). An oxidizer copper (II) ions (Cu²⁺) affected bacterial growth in a concentration-dependent manner (within the range of 0.05 mM to 1 mM) increasing lag phase duration and decreasing specific growth rate. These effects were observed with the wild-type strain ATCC9790 and the atpD mutant strain MS116 (with absent β subunit of F₁ of the F_oF₁ ATPase) both. Also ATPase activity and proton–potassium ions exchange were assessed with and without N,N′-dicyclohexylcarbodiimide (DCCD), inhibitor of the F_oF₁ ATPase. In both cases (DCCD ±), even low Cu²⁺ concentrations had noticeable effect on ATPase activity, but with less visible concentration-dependent manner. Changes in the number of accessible SH-groups were observed with E. hirae ATCC9790 and MS116 membrane vesicles. In both strains Cu²⁺ markedly decreased the number of SH-groups in the presence of K⁺ ions. The addition of ATP increased the amount of accessible SH-groups in ATCC9790 and decreased this number in MS116; Cu²⁺ blocked ATP-installed increase in SH-groups number in ATCC9790. H⁺–K⁺-exchange of bacteria was markedly inhibited by Cu²⁺, but stronger effects were detected together with DCCD. Moreover, discrimination between Cu²⁺ and other bivalent cation—Ni²⁺ was shown. It is suggested that Cu²⁺ ions inhibit E. hirae cell growth by direct affect on the F_oF₁ ATPase leading to conformational changes in this protein complex and decrease in its activity.     

Effects of temperature,photosynthetic photon flux density,photoperiod and O<Subscript>2</Subscript> and CO<Subscript>2</Subscript> concentrations on growth rates of the symbiotic dinoflagellate, <Emphasis Type="Italic">Amphidinium</Emphasis> sp.

Symbiotic dinoflagellates of the species Amphidinium are expected to be pharmaceutically useful microalgae because they produce antitumor macrolides. A microalgae production system with a large number of cells at a high density has been developed for the efficient production of macrolide compounds. In the present study, the effects of culture conditions on the cellular growth rate of dinoflagellates were investigated to determine the optimum culture conditions for obtaining high yields of microalgae. Amphidinium species was cultured under conditions with six temperature levels (21–35°C), six levels of photosynthetic photon flux density (15–70 μmol photons m⁻² s⁻¹), three levels of CO₂ concentration (0.02–0.1%), and three levels of O₂ concentration (0.2–21%). The number of cells cultured in a certain volume of solution was monitored microscopically and the cellular growth rate was expressed as the specific growth rate. The maximum specific growth rate was 0.022 h⁻¹ at a temperature of 26°C and O₂ concentration of 5%, and the specific growth rate was saturated at a CO₂ concentration of 0.05%, a photosynthetic photon flux density of 35 μmol photons m⁻² s⁻¹ and a photoperiod of 12 h day⁻¹ upon increasing each environmental parameter. The results demonstrate that Amphidinium species can multiply efficiently under conditions of relatively low light intensity and low O₂ concentration.     

Phototropic H<Subscript>2</Subscript> production by a newly isolated strain of <Emphasis Type="Italic">Rhodopseudomonas</Emphasis><Emphasis Type="Italic">palustris</Emphasis>

Rhodopseudomonas palustris TN1 was isolated from Songkhla Lake, Thailand. It phototrophically generates H₂ from the predominant volatile fatty acids (VFAs) produced from microbial dark-fermentations of palm oil milling effluent; yields from 20 mM butyrate, acetate and propionate were 4.7, 2.5, and 1.7 mol H₂ mol VFA⁻¹ with light efficiencies of 1.8, 1, and 0.2%, respectively. Optimum conditions were pH 7 and 3000 lux, although production was reduced by only 33% at 1000 lux. CO₂ evolution never exceeded 9 mmol mol VFA⁻¹.     

Effect of nitrogen forms on growth,cell composition and N<Subscript>2</Subscript> fixation of <Emphasis Type="Italic">Cylindrospermopsis raciborskii</Emphasis> in phosphorus-limited chemostat cultures

The aim of this research was to test whether NH₄ ⁺ and NO₃ ⁻ affect the growth, P demand, cell composition and N₂ fixation of Cylindrospermopsis raciborskii under P limitation. Experiments were carried out in P-limited (200 μg l⁻¹ PO₄-P) chemostat cultures of C. raciborskii using an inflowing medium containing either 4,000 μg l⁻¹ NH₄-N, 4,000 μg l⁻¹ NO₃-N or no combined N. The results showed the cellular N:P and C:P ratios of C. raciborskii decreased towards the Redfield ratio with increasing dilution rate (D) due to the alleviation of P limitation. The cellular C:N and carotenoids:chlorophyll-a ratios also decreased with D, predominantly as a result of an increase in the chlorophyll-a and N content. The NH₄ ⁺ and NO₃ ⁻ supply reduced the P maintenance cell quota of C. raciborskii. Consequently, the biomass yield of the N₂-grown culture was significantly lower. The maximum specific growth rate of N₂-grown culture was also the lowest observed. It is suggested that these differences in growth parameters were caused by the P and energy requirement for heterocyte formation, nitrogenase synthesis and N₂ fixation. N₂ fixation was partially inhibited by NO₃ ⁻ and completely inhibited by NH₄ ⁺. It was probably repressed through the high N content of cells at high dissolved N concentrations. These results indicate that C. raciborskii is able to grow faster and maintain a higher biomass under P limitation where a sufficient supply of NH₄ ⁺ or NO₃ ⁻ is maintained. Information gained about the species-specific nutrient and pigment stoichiometry of C. raciborskii could help to access the degree of nutrient limitation in water bodies. Handling editor: Luigi Naselli-Flores