P-limited Cultures Research Articles

Ecological investigations of giant Phaeocystis colonies in Viet Nam: I. Cell abundance and elemental composition.

Phaeocystis globosa is an important bloom-forming marine phytoplankton species that often accumulates to large levels in temperate and tropical waters and has significant impacts on food webs and biogeochemical cycles. It can form "giant" colonies that reach 3 cm in diameter. Microscopic observations, colony elemental composition, and pigment composition were analyzed to assess the characteristics of colonies as a function of colony size. Particulate organic carbon (POC) per unit surface area, colonial cell density, and chlorophyll a per unit surface area all increased with colony size, in contrast to results from temperate waters. Cellular chl a averaged 0.85 pg chl · cell-1. Colonies had both photosynthetic and protective pigments, with fucoxanthin being the dominant accessory pigment. Based on chl a and pigment levels, it appears colonies were acclimated to relatively low irradiances, likely due to their life cycle and the extremely turbulent environment in which they grew. Mucous carbon ranged from 16.2% to 79.2% of the total POC, and mucous carbon per unit surface area increased with colony size, suggesting that the mucous envelope did not thin as the colony grew. Based on elemental composition, nitrogen did not appear to limit growth, but phosphorus:carbon ratios were similar to those of P-limited cultures. Giant colonies represent an extreme response to the environment, but they do not appear to have greatly different characteristics than other tropical strains.

Effects and mechanisms of glyphosate as phosphorus nutrient on element stoichiometry and metabolism in the diatom Phaeodactylum tricornutum.

The ability to utilize dissolved organic phosphorus (DOP) gives phytoplankton competitive advantages in P-limited environments. Our previous research indicates that the diatom Phaeodactylum tricornutum could grow on glyphosate, a DOP with carbon-phosphorus (C-P) bond and an herbicide, as sole P source. However, direct evidence and mechanism of glyphosate utilization are still lacking. In this study, using physiological and isotopic analysis, combined with transcriptomic profiling, we demonstrated the uptake of glyphosate by P. tricornutum and revealed the candidate responsible genes. Our data showed a low efficiency of glyphosate utilization by P. tricornutum, suggesting that glyphosate utilization costs energy and that the alga possessed an herbicide-resistant type of 5-enolpyruvylshikimate-3-phosphate (EPSP) synthase. Compared to the P-limited cultures, the glyphosate-grown P. tricornutum cells up-regulated genes involved in DNA replication, cell growth, transcription, translation, carbon metabolism, and many genes encoding antioxidants. Additionally, cellular C and silicon (Si) increased remarkably while cellular nitrogen (N) declined in the glyphosate-grown P. tricornutum, leading to higher Si:C and Si:N ratios, which corresponded to the up-regulation of genes involved in the C metabolism and Si uptake and the down-regulation of those encoding N uptake. This has the potential to enhance C and Si export to the deep sea when P is limited but phosphonate is available. In sum, our study documented how P. tricornutum could utilize the herbicide glyphosate as P nutrient and how glyphosate utilization may affect the element content and stoichiometry in this diatom, which have important ecological implications in the future ocean.IMPORTANCEGlyphosate is the most widely used herbicide in the world and could be utilized as phosphorus (P) source by some bacteria. Our study first revealed that glyphosate could be transported into Phaeodactylum tricornutum cells for utilization and identified putative genes responsible for glyphosate uptake. This uncovers an alternative strategy of phytoplankton to cope with P deficiency considering phosphonate accounts for about 25% of the total dissolved organic phosphorus (DOP) in the ocean. Additionally, accumulation of carbon (C) and silicon (Si), as well as elevation of Si:C ratio in P. tricornutum cells when grown on glyphosate indicates glyphosate as the source of P nutrient has the potential to result in more C and Si export into the deep ocean. This, along with the differential ability to utilize glyphosate among different species, glyphosate supply in dissolved inorganic phosphorus (DIP)-depleted ecosystems may cause changes in phytoplankton community structure. These insights have implications in evaluating the effects of human activities (use of Roundup) and climate change (potentially reducing DIP supply in sunlit layer) on phytoplankton in the future ocean.

Naphthalene acetic acid and scorbic acid significantly improve neutral lipid accumulation in Chlorella species under nutrient limitations

Nutrient starvation condition has already been exploited as a suitable strategy to induce more lipid production. In this study, nutrient limitation has been coupled with phytohormone or antioxidant supplementation to identify the potential of chemical stimulants to support growth and neutral lipid accumulation. Naphthalene acetic acid (NAA), and Ascorbic acid (AA), at concentrations of 5, 10, and 15 ppm were supplemented to N-limited level 1 (0.25 g NO3-N/L), N-limited level 2 (0.041 gNO3-N/L), and P-limited (0.01 gPO4-P/L) culture media of two Chlorella species to maximize biomass and lipid production. The results show that neutral lipid content increases 70 % when AA is added to the P-limited medium of Chlorella vulgaris IG-R-96 at 15 ppm. At N-limitation level 1 coupled with NAA and AA supplementations at 15 ppm, neutral lipid contents increase 107 % and 128 % in C. vulgaris IG-R-96, and 42 % and 60 % in Chlorella sorokiniana IG-W-96, respectively. When simultaneous N- and P-limitation is applied, neutral lipid content becomes almost double in C. vulgaris IG-R-96 at 10 ppm of NAA supplement. The effects of phytohormone and antioxidant on growth and neutral lipid accumulation is species-dependent and could be more effective when it is combined with nutrient limitation conditions. Stimulant supplementation is significantly effective for neutral lipid accumulation.

Unusual concurrence of P-solubilizing and biocontrol traits under P-limitation in plant-beneficial Pseudomonas aeruginosa P4: insights from in vitro metabolic and gene expression analysis.

Plant-beneficial fluorescent Pseudomonas species with concurrent P-solubilizing and biocontrol traits could have improved rhizospheric survival and efficacy; this rare ability being subject to diverse environmental and endogenous regulations. This study correlates growth patterns, time-course analysis of selected metabolites, non-targeted metabolomics of exometabolites and selected gene expression analysis to elucidate P-limitation-induced physiological shifts enabling co-production of metabolites implied in P-solubilization and biocontrol by P. aeruginosa P4 (P4). P-limited culture supernatants showed enhanced production of selected biocontrol metabolites such as pyocyanin, pyoverdine and pyochelin and IAA while maintaining biomass yield despite reduced growth rate and glucose consumption. Non-targeted exometabolomics further indicated that P-limitation positively impacted pentose phosphate pathway as well as pyruvate, C5-branched dibasic acid and amino acid metabolism. Its correlation with unusually reduced aroC expression and growth phase-dependent changes in the expression of key biosynthetic genes pchA, pchE, pchG, pvdQ and phzM implied a probable regulation of biosynthesis of chorismate-derived secondary metabolites, not neglecting the possibility of multiple factors influencing the gene expression profiles. Similar increase in biocontrol metabolite production was also observed in Artificial Root Exudates (ARE)-grown P4 cultures. While such metabolic flexibility could impart physiological advantage in sustaining P-starvation stress, it manifests as unique coexistence of P-solubilizing and biocontrol abilities.

Nitrogen and phosphorus significantly alter growth, nitrogen fixation, anatoxin-a content, and the transcriptome of the bloom-forming cyanobacterium, Dolichospermum.

While freshwater cyanobacteria are traditionally thought to be limited by the availability of phosphorus (P), fixed nitrogen (N) supply can promote the growth and/or toxin production of some genera. This study characterizes how growth on N2 (control), nitrate (NO3–), ammonium (NH4+), and urea as well as P limitation altered the growth, toxin production, N2 fixation, and gene expression of an anatoxin-a (ATX-A) – producing strain of Dolichospermum sp. 54. The transcriptomes of fixed N and P-limited cultures differed significantly from those of fixed N-deplete, P-replete (control) cultures, while the transcriptomes of P-replete cultures amended with either NH4+ or NO3– were not significantly different relative to those of the control. Growth rates of Dolichospermum (sp. 54) were significantly higher when grown on fixed N relative to without fixed N; growth on NH4+ was also significantly greater than growth on NO3–. NH4+ and urea significantly lowered N2 fixation and nifD gene transcript abundance relative to the control while cultures amended with NO3– exhibited N2 fixation and nifD gene transcript abundance that was not different from the control. Cultures grown on NH4+ exhibited the lowest ATX-A content per cell and lower transcript abundance of genes associated ATX-A synthesis (ana), while the abundance of transcripts of several ana genes were highest under fixed N and P - limited conditions. The significant negative correlation between growth rate and cellular anatoxin quota as well as the significantly higher number of transcripts of ana genes in cultures deprived of fixed N and P relative to P-replete cultures amended with NH4+ suggests ATX-A was being actively synthesized under P limitation. Collectively, these findings indicate that management strategies that do not regulate fixed N loading will leave eutrophic water bodies vulnerable to more intense and toxic (due to increased biomass) blooms of Dolichospermum.

Stoichiometry of Fe, Mn and Co in the marine diazotroph Crocosphaera subtropica ATCC51142 in Fe- and P-limited continuous cultures

We investigated trace element stoichiometries of the nitrogen-fixing marine cyanobacterium Crocosphaera subtropica ATCC51142 under steady-state growth conditions. We utilized exponentially fed batch cultures and varied iron (Fe) concentrations to establish nutrient limitation in C. subtropica growing at a constant growth rate (0.11 d-1). No statistical difference in cell density, chlorophyll a, particulate organic carbon (C), nitrogen (N) and phosphorus (P) were observed between consecutive days after Day 14, and cultures were assumed to be at steady state with respect to growth for the remaining 11 d of the experiment. Cultures were limited by P in the highest Fe treatment (41 nmol l-1) and by Fe in the 2 lower-concentration Fe treatments (1 and 5 nmol l-1). Cell size and in vivo fluorescence changed throughout the experiment in the 1 nmol l-1 Fe treatment, suggesting ongoing acclimation of C. subtropica to our lowest Fe supply. Nevertheless, Fe:C ratios were not significantly different between the Fe treatments, and we calculated an average (±SD) Fe:C ratio of 32 ± 14 µmol mol-1 for growth at 0.11 d-1. Steady-state P-limited cells had lower P quotas, whilst Fe-limited cells had higher manganese (Mn) and cobalt (Co) quotas. We attribute the increase in Mn and Co quotas at low Fe to a competitive effect resulting from changes in the supply ratio of trace elements. Such an effect has implications for variability in elemental stoichiometry in marine phytoplankton, and potential consequences for trace metal uptake and cycling in marine systems.

Distinct Responses of the Nitrogen-Fixing Marine Cyanobacterium Trichodesmium to a Thermally Variable Environment as a Function of Phosphorus Availability.

Surface temperature in the ocean is projected to be elevated and more variable in the future, which will interact with other environmental changes like reduced nutrient supplies. To explore these multiple stressor relationships, we tested the influence of thermal variation on the key marine diazotrophic cyanobacterium Trichodesmium erythraeum GBRTRLI101 as a function of the limiting nutrient phosphorus (P). Two constant temperature treatments represented current winter (22°C) and summer (30°C) mean values. Three variable temperature treatments fluctuated around the constant control values: Mean 22°C, either ± 2°C or ± 4°C; and mean 30°C ± 2°C. Each thermal treatment was grown under both P-replete (10 μmol/L) and P-limiting conditions (0.2 μmol/L). Effects of thermal variability on Trichodesmium were mainly found in the two winter variable temperature treatments (22°C ± 2°C or ± 4°C). P availability affected growth and physiology in all treatments and had significant interactions with temperature. P-replete cultures had higher growth and nitrogen and carbon fixation rates in the 22°C constant control, than in the corresponding variable treatments. However, physiological rates were not different in the P-replete constant and variable treatments at 30°C. In contrast, in P-limited cultures an advantage of constant temperature over variable temperature was not apparent. Phosphorus use efficiencies (PUE, mol N or C fixed h-1 mol cellular P-1) for nitrogen and carbon fixation were significantly elevated under P-limited conditions, and increased with temperature from 22 to 30°C, implying a potential advantage in a future warmer, P-limited environment. Taken together, these results imply that future increasing temperature and greater thermal variability could have significant feedback interactions with the projected intensification of P-limitation of marine N2-fixing cyanobacteria.

Cyanophage Propagation in the Freshwater Cyanobacterium Phormidium Is Constrained by Phosphorus Limitation and Enhanced by Elevated pCO2.

Intensification of human activities has led to changes in the availabilities of CO2 and nutrients in freshwater ecosystems, which may greatly alter the physiological status of phytoplankton. Viruses require hosts for their reproduction and shifts in phytoplankton host physiology through global environmental change may thus affect viral infections as well. Various studies have investigated the impacts of single environmental factors on phytoplankton virus propagation, yet little is known about the impacts of multiple factors, particularly in freshwater systems. We therefore tested the combined effects of phosphorus limitation and elevated pCO2 on the propagation of a cyanophage infecting a freshwater cyanobacterium. To this end, we cultured Phormidium in P-limited chemostats under ambient (400 μatm) and elevated (800 μatm) pCO2 at growth rates of 0.6, 0.3, and 0.05 d-1. Host C:P ratios generally increased with strengthened P-limitation and with elevated pCO2. Upon host steady state conditions, virus growth characteristics were obtained in separate infection assays where hosts were infected by the double-stranded DNA cyanophage PP. Severe P-limitation (host growth 0.05 d-1) led to a 85% decrease in cyanophage production rate and a 73% decrease in burst size compared to the 0.6 d-1 grown P-limited cultures. Elevated pCO2 induced a 96% increase in cyanophage production rate and a 57% increase in burst size, as well as an 85% shorter latent period as compared to ambient pCO2 at the different host growth rates. In addition, elevated pCO2 caused a decrease in the plaquing efficiency and an increase in the abortion percentage for the 0.05 d-1 P-limited treatment, while the plaquing efficiency increased for the 0.6 d-1 P-limited cultures. Together, our results demonstrate interactive effects of elevated pCO2 and P-limitation on cyanophage propagation, and show that viral propagation is generally constrained by P-limitation but enhanced with elevated pCO2. Our findings indicate that global change will likely have a severe impact on virus growth characteristics and thereby on the control of cyanobacterial hosts in freshwater ecosystems.

The role of heterotrophic bacteria in assessing phosphorus stress to Synechocystis sp. PCC6803

Microalgae biofuel production, a possible source of carbon-neutral energy, requires phosphorus (P), a limited resource. This study investigates the relationship between specific growth rate of the cyanobacterium Synechocystis sp. PCC 6803 and P availability. It has been previously suggested, and we show here, that P-limited growth kinetics are well represented by a quota-type model with a single pool of intracellular P. We also demonstrate that the presence of heterotrophic bacteria plays a large role in understanding these kinetics, because the culture’s intracellular P content depends on the level of heterotrophic bacteria. Using batch-growth experiments containing up to a 0.07 biovolume ratio of heterotrophic bacteria to Synechocystis, we found that Synechocystis could grow with intracellular P content down to approximately 0.5 mg P g−1 dry weight biomass, while heterotrophic bacteria maintained roughly 20 mg P g−1 dry weight. Thus, a small fraction of heterotrophic bacteria in a microalgal culture can dramatically increase the apparent content of P in the biomass, which affects how to assess P-stress to a P-limited culture.

Recycling harvest water to cultivate Chlorella zofingiensis under nutrient limitation for biodiesel production

Harvest water recycling for Chlorella zofingiensis re-cultivation under nutrient limitation was investigated. Using 100% harvest water, four cultures were prepared: Full medium, P-limited medium, N-limited medium and N- and P-limited medium, while another full medium was also prepared using 50% harvest water. The results showed that the specific growth rate and biomass productivity ranged from 0.289 to 0.403 day(-1) and 86.30 to 266.66 mg L(-1) day(-1), respectively. Nutrient-limited cultures witnessed much higher lipid content (41.21-46.21% of dry weight) than nutrient-full cultures (26% of dry weight). The N- and P-limited medium observed the highest FAME yield at 10.95% of dry weight, while the N-limited culture and P-limited culture shared the highest biodiesel productivity at 20.66 and 19.91 mg L(-1) day(-1), respectively. The experiment on harvest water recycling times demonstrated that 100% of the harvest water could be recycled twice with the addition of sufficient nutrients.

Increased toxicity of Karenia brevis during phosphate limited growth: ecological and evolutionary implications.

Karenia brevis is the dominant toxic red tide algal species in the Gulf of Mexico. It produces potent neurotoxins (brevetoxins [PbTxs]), which negatively impact human and animal health, local economies, and ecosystem function. Field measurements have shown that cellular brevetoxin contents vary from 1–68 pg/cell but the source of this variability is uncertain. Increases in cellular toxicity caused by nutrient-limitation and inter-strain differences have been observed in many algal species. This study examined the effect of P-limitation of growth rate on cellular toxin concentrations in five Karenia brevis strains from different geographic locations. Phosphorous was selected because of evidence for regional P-limitation of algal growth in the Gulf of Mexico. Depending on the isolate, P-limited cells had 2.3- to 7.3-fold higher PbTx per cell than P-replete cells. The percent of cellular carbon associated with brevetoxins (%C-PbTx) was ∼ 0.7 to 2.1% in P-replete cells, but increased to 1.6–5% under P-limitation. Because PbTxs are potent anti-grazing compounds, this increased investment in PbTxs should enhance cellular survival during periods of nutrient-limited growth. The %C-PbTx was inversely related to the specific growth rate in both the nutrient-replete and P-limited cultures of all strains. This inverse relationship is consistent with an evolutionary tradeoff between carbon investment in PbTxs and other grazing defenses, and C investment in growth and reproduction. In aquatic environments where nutrient supply and grazing pressure often vary on different temporal and spatial scales, this tradeoff would be selectively advantageous as it would result in increased net population growth rates. The variation in PbTx/cell values observed in this study can account for the range of values observed in the field, including the highest values, which are not observed under N-limitation. These results suggest P-limitation is an important factor regulating cellular toxicity and adverse impacts during at least some K. brevis blooms.

Photosynthetic efficiency, cell volume, and elemental stoichiometric ratios in Thalassirosira weissflogii under phosphorus limitation

Nutrient limitation is known to inhibit growth and metabolism and to alter elemental stoichiometric ratios in phytoplankton. In this study, physiological changes in Thalassirosira weissflogii were measured under different dissolved inorganic phosphate (DIP) regimes in semi-continuous cultures to revisit the utility of the Redfield ratio for assessing nutrient limitation. The results showed that cell size increased with decreasing DIP availability. In the P-depleted treatment (f/2-P) the cell size was 1.48 times larger than that in the P-limited (f/100) treatment and 2.67 times larger than that in the P-saturated treatment (f/2 and f/10). The fucoxanthin to chlorophyll a ratio (Fuco/chl a) was relatively stable (about 0.3) in P-saturated cultures and was 10 times higher than that in P-limited and P-depleted cultures. During the experimental period, the photosynthetic efficiency index, ΔF/Fm′, was relatively stable at ∼0.50 in the P-saturated cultures, but quickly declined with decreasing DIP availability. Although cellular P content showed a significant difference between the P-saturated culture (1.6 pg/cell) and the P-limited culture (0.7 pg/cell), the N/P ratio in T. weissflogii did not show a trend with DIP availability and fluctuated slightly around 25. Our results suggest that cell division in T. weissflogii is not strictly size-gated but is probably regulated by a biochemical, and hence, an elemental stoichiometric ratio threshold, and that deviation of the cellular N/P ratio from the Redfield ratio is not a reliable indicator of algal nutrient stress.

Effects of changing pCO2 and phosphate availability on domoic acid production and physiology of the marine harmful bloom diatom Pseudo‐nitzschia multiseries

Some members of the diatom genus Pseudo‐nitzschia produce the toxin domoic acid (DA), which through trophic transfer causes mass mortalities of wildlife, shellfish harvesting closures, and risks to human health. Nutrient and micronutrient limitation have been shown to regulate DA production. This study tested the hypothesis that changing partial pressure of CO2 (pCO2) can interact with nutrient limitation to help determine cellular DA levels, an environmentally relevant issue in light of current increases in atmospheric pCO2. Cultures of the toxic species Pseudo‐nitzschia multiseries were incubated using semicontinuous methods under a matrix of three pCO2 conditions: ∼ 22 Pa (220 ppm), ∼ 41 Pa (400 ppm), and ~ 74 Pa (730 ppm), and two phosphate concentrations: 20 μmol L−1, P‐replete; and 0.5 μmol L−1, P‐limited. DA production was regulated by both pCO2 and phosphate availability. DA concentrations were 30–50 times higher in P‐limited cultures compared to P‐replete ones, at the same pCO2 levels. Increasing CO2 levels stimulated DA production under both nutrient conditions, but especially in P‐limited cultures, where DA levels increased approximately four times over the pCO2 range examined. Growth rates, primary productivity, photosynthesis vs. irradiance parameters, and cellular elemental ratios also responded interactively to the availability of both CO2 and phosphate. Our results raise the possibility that growth rates and toxicity of the diatom Pseudo‐nitzschia multiseries could increase substantially in the future high‐CO2 ocean, suggesting a potentially escalating negative effect of this harmful algal bloom species on the future marine environment.

Genome-wide transcriptomic analysis of the response to nitrogen limitation in Streptomyces coelicolor A3(2)

BackgroundThe present study represents a genome-wide transcriptomic analysis of the response of the model streptomycete Streptomyces coelicolor A3(2) M145 to fermentor culture in Modified Evans Media limited, respectively, for nitrogen, phosphate and carbon undertaken as part of the ActinoGEN consortium to provide a publicly available reference microarray dataset.FindingsA microarray dataset using samples from two replicate cultures for each nutrient limitation was generated. In this report our analysis has focused on the genes which are significantly differentially expressed, as determined by Rank Products Analysis, between samples from matched time points correlated by growth phase for the three pairs of differently limited culture datasets. With a few exceptions, genes are only significantly differentially expressed between the N6/N7 time points and their corresponding time points in the C and P-limited cultures, with the vast majority of the differentially expressed genes being more highly expressed in the N-limited cultures. Our analysis of these genes indicated expression of several members of the GlnR regulon are induced upon nitrogen limitation, as assayed for by [NH4+] measurements, and we are able to identify several additional genes not present in the GlnR regulon whose expression is induced in response to nitrogen limitation. We also note SCO3327 which encodes a small protein (32 amino acid residues) unusually rich in the basic amino acids lysine (31.25%) and arginine (25%) is significantly differentially expressed in the nitrogen limited cultures. Additionally, we investigate the expression of known members of the GlnR regulon and the relationship between gene organization and expression for the SCO2486-SCO2487 and SCO5583-SCO5585 operons.ConclusionsWe provide a list of genes whose expression is differentially expressed in low nitrogen culture conditions, including a putative nitrogen storage protein encoded by SCO3327. Our list includes several genes whose expression patterns are similar to up-regulated members of the GlnR regulon and are induced in response to nitrogen limitation. These genes represent likely targets for future studies into the nitrogen starvation response in Streptomyces coelicolor.

CO2 and phosphate availability control the toxicity of the harmful bloom dinoflagellate Karlodinium veneficum

AME Aquatic Microbial Ecology Contact the journal Facebook Twitter RSS Mailing List Subscribe to our mailing list via Mailchimp HomeLatest VolumeAbout the JournalEditorsSpecials AME 59:55-65 (2010) - DOI: https://doi.org/10.3354/ame01396 CO2 and phosphate availability control the toxicity of the harmful bloom dinoflagellate Karlodinium veneficum Fei-Xue Fu1,*, Allen R. Place2, Nathan S. Garcia1, David A Hutchins1 1Department of Biological Sciences, University of Southern California, 3616 Trousdale Parkway, Los Angeles, California 90089, USA 2Center of Marine Biotechnology, University of Maryland Biotechnology Institute, Baltimore, Maryland 21202, USA *Email: ffu@usc.edu ABSTRACT: We demonstrated that the toxicity of the harmful bloom dinoflagellate Karlodinium veneficum is regulated by both CO2 concentrations and phosphate availability. Semi-continuous cultures were grown in a factorial experiment under all combinations of 3 CO2 levels (230, 430 and 745 ppm) and 2 phosphate conditions (0.5 and 20 µM). After steady-state acclimation was achieved, karlotoxin cellular quotas and growth rates were determined in all 6 treatments. This strain produced both types of karlotoxin, KmTx-1 and KmTx-2. Chlorophyll a-normalized production of the 2 types of karlotoxins was much higher in P-limited cultures compared with P-replete ones under the same CO2 conditions. Increasing CO2 strongly stimulated production of KmTx-1 and decreased production of KmTx-2 in both treatments, but especially in P-limited cultures. Because the KmTx-1 toxin is an order of magnitude more potent than KmTx-2, total cellular toxicity was increased dramatically at high pCO2, particularly in P-limited cultures. Specific growth rates were accelerated by enriched CO2 in P-replete cultures, but not in P-limited treatments. Growth rates or toxicity of K. veneficum could increase substantially in the future with high CO2 levels in the ocean, depending on P availability, and so interactions between rising CO2 and eutrophication could cause major shifts in present day patterns of harmful algal toxin production. These results suggest that over the coming decades, rising CO2 could substantially increase karlotoxin damage to food webs in the often P-limited estuaries where Karlodinium blooms occur. KEY WORDS: Harmful algae · Karlodinium veneficum · Karlotoxin · CO2 · Phosphate · Dinoflagellates · Ocean acidification · Algal toxin Full text in pdf format PreviousNextCite this article as: Fu FX, Place AR, Garcia NS, Hutchins DA (2010) CO2 and phosphate availability control the toxicity of the harmful bloom dinoflagellate Karlodinium veneficum. Aquat Microb Ecol 59:55-65. https://doi.org/10.3354/ame01396 Export citation RSS - Facebook - Tweet - linkedIn Cited by Published in AME Vol. 59, No. 1. Online publication date: March 11, 2010 Print ISSN: 0948-3055; Online ISSN: 1616-1564 Copyright © 2010 Inter-Research.

Relationship between phosphate affinities and cell size and shape in various bacteria and phytoplankton

Substrate affinity expresses the ability of an osmotroph organism to compete for a sub- strate at permanently low external concentrations and is thus a central parameter in conceptual and mathematical models of aquatic food webs. Assuming diffusion transport in the surrounding medium to be the limiting process at low external substrate concentrations, the theoretical maximum affinity (α max ) and its dependence on cell size and shape for a given osmotroph organism can be calculated from Fick's law of diffusion in combination with knowledge of the amount of substrate required to form a new cell. For a non-respired substrate, the actual affinity (α) can also be expressed as the bio- mass-specific turnover rate of the substrate, α = (TB) -1 . Combining a measure of biomass (B), with determination of substrate turnover time (T), the affinity can thus be determined experimentally. We used this approach to compare measured with theoretical maximum affinities for phosphate in labo- ratory cultures of osmotrophic microorganisms. For bacteria and autotrophic flagellates, we found relatively good agreement between experimental and theoretical maximum values, suggesting that diffusion limitation is actually approached in P-limited cultures. Assuming P-free vacuoles, the the- ory predicts diatom affinities to exceed that of similarly sized flagellates. This prediction is consistent with our experimental observations. Previous reports of diatoms being unsuccessful under P-limited conditions may therefore need a more complex explanation than lack of competitive ability in diatoms.

Growth and toxin production of tropical Alexandrium minutum Halim (Dinophyceae) under various nitrogen to phosphorus ratios

Effects of nitrogen to phosphorous (N/P) ratios of two nitrogen sources (nitrate and ammonium) on growth and toxin production of a tropical estuarine dinoflagellate, Alexandrium minutum Halim, were examined using a strain isolated from a bloom at Tumpat Estuary, Malaysia in September 2001. Experiments were carried out in batch cultures, using either nitrate (N-NO3) or ammonium (N-NH4) as the nitrogen source at a constant amount, and with initial N/P ratios ranging from 5 to 500. Cell density, residual N and P in the medium, cellular toxin quota (Q t), and toxin composition were analyzed throughout the growths. Our results showed that cell densities and growth rates of A. minutum were severely suppressed under high N/P ratios (>100) in both N-NO3 and N-NH4 treatments. Cells tended to be larger at lower growth rate and P-limited cultures. Toxin profile was relatively constant throughout the experiments, with GTX4/GTX1 as the dominant toxin congeners. Cellular toxin quota (Q t) increased with elevated N/P ratios in both N-NO3 and N-NH4 treatments. Toxin production rate, R tox, however was enhanced in N-NH4-grown cultures when P was limited, but showed no difference between N-NO3- and N-NH4-grown cultures when P was replete. Our results clearly showed that N/P ratios as well as the nitrogen compounds not only affected the growth of A. minutum, but also the cellular toxin quota and its toxin production rate.

Alkaline phosphatase activity of marine bacteria studied with ELF 97 substrate: success and limits in the P-limited Mediterranean Sea

The fluorogenic substrate Enzyme-Labeled Fluorescence 97 (ELF-P) is hydrolyzed by the P-cleaving enzyme phosphatase, producing ELF 97 alcohol (ELFA), a fluorescent-insoluble product. This reaction is used for monitoring phosphatase activity at the single-cell level. Most frequently, ELF-P has been used to determine the P-limitation status of microphytoplankton, but rarely of heterotrophic bacteria. We incubated ELF-P on filters to monitor marine bacterial cultures and oligotrophic Mediterranean Sea samples. Results were compared to classical measurements of bulk alkaline phosphatase activity using the fluorogenic substrate 4-methylumbelliferyl phosphate (MUF-P). A high percentage of the cultured cells were labeled with ELFA (the ratio of ELFA spots to total DAPI counts in P-limited cultures ranged from 26 to 100%, depending on the strain). In contrast, this ratio never exceeded 0.01 % in Mediterranean samples, even when P was demonstrated to be a significant limiting factor. This protocol is useful for application on cruises and with cultures, but was not sufficiently sensitive to detect P-stressed bacterial cells in oligotrophic marine environments.

Changes in Morphology and Elemental Composition of Vibrio splendidus Along a Gradient from Carbon-limited to Phosphate-limited Growth

We examined morphology, elemental composition (C, N, P), and orthophosphate-uptake efficiency in the marine heterotrophic bacterium Vibrio splendidus grown in continuous cultures. Eight chemostats were arranged along a gradient of increasing glucose concentrations in the reservoirs, shifting the limiting factor from glucose to phosphate. The content of carbon, nitrogen, and phosphorus was measured in individual cells by x-ray microanalysis using a transmission electron microscope (TEM). Cell volumes (V) were estimated from length and width measurements of unfixed, air-dried cells in TEM. There was a transition from coccoid cells in C-limited cultures toward rod-shaped cells in P-limited cultures. Cells in P-limited cultures with free glucose in the media were significantly larger than cells in glucose-depleted cultures (P < 0.0001). We found functional allometry between cellular C-, N-, and P content (in femtograms) and V (in cubic micrometers) in V. splendidus (C = 224 x V(0.89), N = 52.5 x V(0.80), P = 2 x V(0.65)); i.e., larger bacteria had less elemental C, N, and P per V than smaller cells, and also less P relative to C. Biomass-specific affinity for orthophosphate uptake in large P-limited V. splendidus approached theoretical maxima predicted for uptake limited by molecular diffusion toward the cells. Comparing these theoretical values to respective values for the smaller, coccoid, C-limited V. splendidus indicated, contrary to the traditional view, that large size did not represent a trade-off when competing for the non-C-limiting nutrients.

Nutrition and growth kinetics in nitrogen- and phosphorus-limited cultures of the novel red tide flagellate Chattonella ovata (Raphidophyceae)

In the summer of 2004, a harmful bloom caused by Chattonella ovata (Raphidophyceae) occurred over almost the entire area of the Seto Inland Sea and caused fishery damages. This incident was the first record of a bloom and damage to the fisheries caused by this species in Japanese waters. In order to elucidate the mechanism of the bloom outbreak, we examined the nutrition and the growth kinetics in nitrogen (N)- and phosphorus (P)-limited semi-continuous cultures of this species. Inorganic N compounds, such as nitrate, nitrite, and ammonium, were found to be good nitrogen sources for the growth of C. ovata, while organic nitrogen (urea and uric acid) was not utilized. This species was capable of using ATP, ADP and inorganic phosphorus compounds, but did not utilize phosphate monoesters as a sole P source. Under both N-limited and P-limited steady state conditions, the growth rate as a function of cell nitrogen and phosphorus quota, respectively, followed the Droop equation. Kinetic parameters μ m (maximum growth rate) and k q (minimum cell quota) obtained for N- and P-limited cultures were 0.79 day −1 and 5.5 pmol N cell −1 and 0.86 day −1 and 0.48 pmol P cell −1, respectively. The minimum cell quotas were 23–30% lower than those of C. antiqua. The nutrient availability and kinetic parameters of C. ovata are compared with other harmful algae and the ecological implications of these characteristics discussed.