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During a 2-year study of planktonic nitrogen (N) nutrition, temporal variability of (1) ambient nutrient concentrations; (2) uptake rates of ammonium (NH4+), nitrate (NO3−), nitrite (NO2−), urea, and amino acids (AA) in three size fractions (> GF/F, > 5 μm, and 5–0.2 μm); (3) NH4+ regeneration and NO3− regeneration (nitrification); and (4) an unexpected bloom of Alexandrium monilatum were examined. Dissolved organic N (DON) was the most abundant form of fixed N. High concentrations of NH4+ and NO2− were detected during the late summer and fall, reaching maximums of 9.9 and 7.6 μmol N L−1, respectively. The highest uptake rates were for NH4+ at all stations, size fractions, and seasons sampled and ranged from 34 to 80% of total absolute N uptake. The magnitude of uptake rates in the > GF/F fraction generally followed the pattern of NH4+ > NO3− > urea > AA > NO2− with some exceptions when urea uptake rates were higher than NO3−. Rates of NH4+ regeneration and nitrification often exceeded uptake rates, indicating autochthonous pathways for nutrient loading. Exceptionally high dinoflagellate biomass was found in late summer and corresponded with harmful algal blooms. Kinetic curves measured during an A. monilatum bloom showed high Vmax (33.7 ± 2.7 × 10−3 h−1) and high Ks (7.3 μmol N L−1) for NH4+ suggesting that it can rapidly utilize high concentrations when available but may be outcompeted by other phytoplankton when concentrations of NH4+ are low. However, A. monilatum demonstrated that it is capable of using a diverse suite of N substrates, giving it a potential competitive advantage under diverse nutrient conditions.

Significance Phytoplankton contribute to the Southern Ocean’s (SO) ability to absorb atmospheric CO2 and shape the stoichiometry of northward macronutrient delivery. Climate change is altering the SO environment, yet we know little about how resident phytoplankton will react to these changes. Here, we studied a natural SO community and compared responses of two prevalent, bloom-forming diatom groups to changes in temperature and iron that are projected to occur by 2100 to 2300. We found that one group, Pseudo-nitzschia, grows better under warmer low-iron conditions by managing cellular iron demand and efficiently increasing photosynthetic capacity. This ability to grow and draw down nutrients in the face of warming, regardless of iron availability, has major implications for ocean ecosystems and global nutrient cycles., The Southern Ocean (SO) harbors some of the most intense phytoplankton blooms on Earth. Changes in temperature and iron availability are expected to alter the intensity of SO phytoplankton blooms, but little is known about how these changes will influence community composition and downstream biogeochemical processes. We performed light-saturated experimental manipulations on surface ocean microbial communities from McMurdo Sound in the Ross Sea to examine the effects of increased iron availability (+2 nM) and warming (+3 and +6 °C) on nutrient uptake, as well as the growth and transcriptional responses of two dominant diatoms, Fragilariopsis and Pseudo-nitzschia. We found that community nutrient uptake and primary productivity were elevated under both warming conditions without iron addition (relative to ambient −0.5 °C). This effect was greater than additive under concurrent iron addition and warming. Pseudo-nitzschia became more abundant under warming without added iron (especially at 6 °C), while Fragilariopsis only became more abundant under warming in the iron-added treatments. We attribute the apparent advantage Pseudo-nitzschia shows under warming to up-regulation of iron-conserving photosynthetic processes, utilization of iron-economic nitrogen assimilation mechanisms, and increased iron uptake and storage. These data identify important molecular and physiological differences between dominant diatom groups and add to the growing body of evidence for Pseudo-nitzschia’s increasingly important role in warming SO ecosystems. This study also suggests that temperature-driven shifts in SO phytoplankton assemblages may increase utilization of the vast pool of excess nutrients in iron-limited SO surface waters and thereby influence global nutrient distribution and carbon cycling.

Effluent discharged from water reclamation facilities (WRFs) contains dissolved organic nitrogen, termed effluent dissolved organic nitrogen (EDON), that subsequently enters coastal waterways. It is still unclear at what rate EDON can be taken up by microbial communities relative to other nitrogen (N) substrates. Bench-scale sequencing batch reactors (SBRs), used to mimic WRFs, were supplied with 15N-labeled ammonium (15NH4+) to produce 15N-labeled EDON (EDO15N) that was subsequently used to measure uptake rates along a salinity gradient of the York River, Virginia, USA, in the spring and summer. Although NH4+ dominated influent N pools, only a small fraction (4.1%) of EDON was produced from NH4+ microbial assimilation in biological treatment processes. When added as a short-term (4-h) tracer, the EDO15N was taken up by estuarine microbes at rates 0.01–0.434 μmol N L−1 h−1, which are similar to rates of NH4+ and nitrate uptake. When added to 48-h bioassays, EDON stimulated phytoplankton growth more at the lower salinity (0–8‰) sites (8.5–13.8 μg Chl a L−1) than at the higher salinity (20‰) site (up to 0.4 μmol Chl a L−1). The microbes in the 0.7–5 μm size fraction had significantly higher EDO15N uptake rates than the larger size fraction (e.g., > 5 μm, p

The Southern Ocean (SO) harbours some of the most intense phytoplankton blooms on Earth. Changes in temperature and iron availability are expected to alter the intensity of SO phytoplankton blooms, but little is known about how environmental change will influence community composition and downstream biogeochemical processes. We performed experimental manipulations on surface ocean microbial communities from McMurdo Sound in the Ross Sea, with and without iron addition, at −0.5 °C, 3 °C, and 6 °C. We then examined nutrient uptake patterns as well as the growth and molecular responses of two dominant diatoms, Fragilariopsis and Pseudo-nitzschia, to these conditions. We found that nitrate uptake and primary productivity were elevated at increased temperature in the absence of iron addition, and were even greater at high temperature with added iron. Pseudo-nitzschia became more abundant under increased temperature without added iron, while Fragilariopsis required additional iron to benefit from warming. We attribute the apparent advantage Pseudo-nitzschia shows under warming to upregulation of iron-conserving photosynthetic processes, utilization of iron-economic nitrogen assimilation mechanisms, and increased iron uptake and storage. These data identify important molecular and physiological differences between dominant diatom groups and add to the growing body of evidence for Pseudo-nitzschia’s increasingly important role in warming SO ecosystems. This study also suggests that temperature-driven shifts in SO phytoplankton assemblages may increase utilization of the vast pool of excess nutrients in iron-limited SO surface waters, and thereby influence global nutrient distributions and carbon cycle.Significance StatementPhytoplankton assemblages contribute to the Southern Ocean’s ability to absorb atmospheric CO2, form the base of marine food webs, and shape the global distribution of macronutrients. Anthropogenic climate change is altering the SO environment, yet we do not fully understand how resident phytoplankton will react to this change. By comparing the responses of two prominent SO diatom groups to changes in temperature and iron in a natural community, we find that one group, Pseudo-nitzschia, grows better under warmer low-iron conditions by managing cellular iron demand and efficiently increasing photosynthetic capacity. This ability to grow and draw down nutrients in the face of warming, regardless of iron availability, may have major implications for ocean ecosystems and global nutrient and carbon cycles.

Dinitrogen (N2) fixation is the source of all biologically available nitrogen on earth, and its presence or absence impacts net primary production and global biogeochemical cycles. Here, we report rates of 3.5–17.2 nmol N L−1 d−1 in the ice‐free coastal Alaskan Arctic to show that N2 fixation in the Arctic Ocean may be an important source of nitrogen to a seasonally nitrogen‐limited system. If widespread in surface waters over ice‐free shelves throughout the Arctic, N2 fixation could contribute up to 3.5 Tg N yr−1 to the Arctic nitrogen budget. At these rates, N2 fixation occurring in ice‐free summer waters would offset up to 27.1% of the Arctic denitrification deficit and contribute an additional 2.7% to N2 fixation globally, making it an important consideration in the current debate of whether nitrogen in the global ocean is in steady state. Additional investigations of high‐latitude marine diazotrophic physiology are required to refine these N2 fixation estimates.

Here, we present the first study to investigate the seasonal importance of amino acid-nitrogen (N) to Arctic near shore microbial communities. We measured primary productivity and the uptake of ammonium, nitrate, urea, and amino acids in two size fractions (> 3 μm and approximately 0.7–3 μm), as well as ammonium regeneration and nitrification using 15N and 13C tracer approaches in the near-shore waters of the Chukchi Sea, during January, April, and August for two consecutive years. At discrete depths, nitrate comprised 46–78% of the total dissolved N pool during January and April but only 2–6% during August. Dissolved organic N (DON) concentrations increased between January and August though the carbon (C) : N (mol : mol) of the DON pool declined. Of the substrates tested, amino acids supported the bulk of both N and C nutrition in both size fractions during January and April (ice-covered). Urea generally had the lowest uptake rate under ice-covered conditions; uptake of urea-C was only detectable in August. Though previous Arctic studies focused largely on nitrate, we found nitrate uptake was generally lower than other substrates tested. The sharp decline in nitrate concentration between April and August, however, indicates a drawdown of nitrate during that period. Rates of ammonium uptake were highest in August, when it was the dominant N substrate used. During all sample periods, rates of ammonium regeneration were sufficient to supply ammonium demand. Rates of nitrification varied between sample periods, however, with much higher rates seen in January and April.

The Arctic is warming at a rate nearly twice the global average, leading to thawing permafrost, increased coastal erosion, and enhanced delivery of riverine terrestrially-derived dissolved organic matter (tDOM) to coastal waters. This humic-rich tDOM has the ability to attenuate light required for photosynthesis and stimulate heterotrophic growth by supplying a source of labile organic carbon. Due to tDOM's high carbon to nitrogen (C : N) ratio, additional nitrogen is required for microorganisms to utilize this excess carbon for growth, thus exacerbating competition between autotrophs and heterotrophs for limiting nutrients and potentially reducing primary production. The effect of Arctic tDOM additions on nitrate uptake by two microplankton size fractions in the coastal Chukchi Sea was quantified using 15N tracer methods. To assess the biogeochemical vs. spectral impacts of tDOM, the uptake incubations were amended with either tDOM or light attenuating films that mimic light absorption by the tDOM. Nitrate uptake and primary production rates in the larger, predominantly phytoplankton, size fraction generally decreased with increasing tDOM additions. The change in light attenuation alone accounted for a ∼ 50% reduction in nitrate uptake. Responses in the smaller size fraction varied seasonally with tDOM additions stimulating uptake in spring and suppressing it in summer. The largest variation in summer nitrate uptake can be explained by the shared effect of biogeochemistry and light attenuation. Therefore, large increases in tDOM delivery currently occurring and predicted to increase in the coastal Arctic, could reduce primary production, broadly impact nitrogen and carbon cycling, and affect higher trophic levels.

Southern Ocean primary productivity plays a key role in global ocean biogeochemistry and climate. At the Southern Ocean sea ice edge in coastal McMurdo Sound, we observed simultaneous cobalamin and iron limitation of surface water phytoplankton communities in late Austral summer. Cobalamin is produced only by bacteria and archaea, suggesting phytoplankton-bacterial interactions must play a role in this limitation. To characterize these interactions and investigate the molecular basis of multiple nutrient limitation, we examined transitions in global gene expression over short time scales, induced by shifts in micronutrient availability. Diatoms, the dominant primary producers, exhibited transcriptional patterns indicative of co-occurring iron and cobalamin deprivation. The major contributor to cobalamin biosynthesis gene expression was a gammaproteobacterial population, Oceanospirillaceae ASP10-02a. This group also contributed significantly to metagenomic cobalamin biosynthesis gene abundance throughout Southern Ocean surface waters. Oceanospirillaceae ASP10-02a displayed elevated expression of organic matter acquisition and cell surface attachment-related genes, consistent with a mutualistic relationship in which they are dependent on phytoplankton growth to fuel cobalamin production. Separate bacterial groups, including Methylophaga, appeared to rely on phytoplankton for carbon and energy sources, but displayed gene expression patterns consistent with iron and cobalamin deprivation. This suggests they also compete with phytoplankton and are important cobalamin consumers. Expression patterns of siderophore- related genes offer evidence for bacterial influences on iron availability as well. The nature and degree of this episodic colimitation appear to be mediated by a series of phytoplankton-bacterial interactions in both positive and negative feedback loops.

The coastal Chukchi Sea has a particularly productive landfast ice ecosystem but is currently lacking direct measurements of nitrogen (N) cycling within the ice. Using stable isotopic tracers, we measured uptake and regeneration of ammonium and nitrate, along with primary and secondary production at three depths within landfast sea ice in the Chukchi Sea near Barrow, Alaska, during April 2011 and January 2012. These are the first data of N uptake and regeneration rates in landfast Arctic sea ice during spring and winter. Both inorganic and organic nutrient concentrations in the ice were generally higher than the water column, with the exception of phosphate, which may have limited production in certain sections of ice. Primary production at all ice depths was higher than the water column during April, but below the detection limit in January. Bacterial production in the bottom ice (0–10 cm from the ice–water interface) was consistently higher than the water column across seasons. Absolute uptake of ammonium was highest in the bottom ice and higher than absolute uptake of nitrate at all depth horizons except in the upper ice (30–40 cm from ice–water interface) during January. While N uptake rates in the ice were higher than the water column, nitrification rates were lower. Regeneration of ammonium and nitrate far exceeded uptake within the ice. The magnitude of N uptake and regeneration rates in landfast ice highlights the importance of the biological sea ice community to the region.

Biogeochemical rate processes in the Arctic are not currently well constrained, and there is very limited information on how rates may change as the region warms. Here we present data on the sensitivity of ammonium (NH4+) uptake and nitrification rates to short-term warming. Samples were collected from the Chukchi Sea off the coast of Barrow, Alaska, during winter, spring, and summer and incubated for 24 h in the dark with additions of 15NH4+ at −1.5, 6, 13, and 20°C. Rates of NH4+ uptake and nitrification were measured in conjunction with bacterial production. In all seasons, NH4+ uptake rates were highest at temperatures similar to current summertime conditions but dropped off with increased warming, indicative of psychrophilic (i.e., cold-loving) microbial communities. In contrast, nitrification rates were less sensitive to temperature and were higher in winter and spring compared to summer. These findings suggest that as the Arctic coastal ecosystem continues to warm, NH4+ assimilation may become increasingly important, relative to nitrification, although the magnitude of NH4+ assimilation would be still be lower than nitrification.

How Arctic climate change might translate into alterations of biogeochemical cycles of carbon (C) and nitrogen (N) with respect to inorganic and organic N utilization is not well understood. This study combined 15 N uptake rate measurements for ammonium, nitrate, and urea with 15 N- and 13 C-based DNA stable-isotope probing (SIP). The objective was to identify active bacterial and archeal plankton and their role in N and C uptake during the Arctic summer and winter seasons. We hypothesized that bacteria and archaea would successfully compete for nitrate and urea during the Arctic winter but not during the summer, when phytoplankton dominate the uptake of these nitrogen sources. Samples were collected at a coastal station near Barrow, AK, during August and January. During both seasons, ammonium uptake rates were greater than those for nitrate or urea, and nitrate uptake rates remained lower than those for ammonium or urea. SIP experiments indicated a strong seasonal shift of bacterial and archaeal N utilization from ammonium during the summer to urea during the winter but did not support a similar seasonal pattern of nitrate utilization. Analysis of 16S rRNA gene sequences obtained from each SIP fraction implicated marine group I Crenarchaeota (MGIC) as well as Betaproteobacteria , Firmicutes , SAR11, and SAR324 in N uptake from urea during the winter. Similarly, 13 C SIP data suggested dark carbon fixation for MGIC, as well as for several proteobacterial lineages and the Firmicutes . These data are consistent with urea-fueled nitrification by polar archaea and bacteria, which may be advantageous under dark conditions.

Identification and quantification of the nutrient sources supporting large, extended duration Karenia brevis blooms on the West Florida Shelf (WFS) in the eastern Gulf of Mexico are critical steps for effective bloom management and mitigation. Previous research had identified multiple (>12) potential nutrient sources available to K. brevis blooms on the WFS, which vary with bloom stage, location, biomass and bloom toxicity. This current study newly identified and quantified additional nitrogen (N) sources including water column nitrification, photochemical nutrient production, pelagic unicell N2 fixation by diazotrophs other than the colonial cyanobacterium Trichodesmium, and remineralization from seasonal Trichodesmium biomass decay and microzooplankton grazing (and estimated regeneration). Newly identified phosphorus (P) sources include remineralization from Trichodesmium biomass decay and microzooplankton grazing. In estuarine environments, benthic nutrient flux, mixotrophic consumption of picoplankton, nutrient release from zooplankton and microzooplankton grazing, photochemical nutrient production, and nitrification all can contribute up to 100% of the N and/or P requirements of small ( Given the complexity of K. brevis bloom dynamics, the multiple available nutrient sources on the WFS and the importance of regenerated N forms in supporting blooms, efforts to reduce potentially controllable nearshore nutrient inputs should be undertaken with the understanding that while they may lead to enhanced coastal water quality, they may not have an immediate impact on the frequency or magnitude of nearshore K. brevis blooms. Additionally, time lags in ecosystem responses or differences in the time scales on which various process operate may require multi-year assessments to determine how effective management practices are in relation to K. brevis blooms. Timely red tide related monitoring products that allow for effective focusing of monitoring needs for short-term prediction of impacts and targeted communication of scientific results to the public and stakeholders, remains the most effective means of K. brevis management.

To investigate nutrient release from decaying fish carcasses as a potential source fueling blooms of the toxic dinoflagellate, Karenia brevis (C.C. Davis) G. Hansen & O. Moestrup this study evaluated the type and quantity of nutrients (nitrogen (N), phosphorus (P), and carbon (C)) released from decaying fish using both wet chemistry and compound specific (Fourier transform ion cyclotron resonance mass spectrometry, FT-ICR MS) techniques. A 3 day bioassay experiment was then conducted to examine how eutrophic (Charlotte Harbor, CH) and oligotrophic (Blue Water, BW) microbial communities in the eastern Gulf of Mexico (GoM) respond to fish-derived nutrients (FDN) relative to other in situ N sources. Decaying fish released a suite of compounds, primarily in the form of ammonium (NH 4 + ); release rates ranged from 37.7 ± 12.6 to 102.3 ± 4.7 μmol N L −1 g −1 wet wt. d −1 . Phosphate release was 13.9 ± 2.8 to 21.4 ± 2.9 μmol P L −1 g −1 wet wt. d −1 . A total of 3515 organic compounds were produced, which were dominated by lipid and protein-like compounds. In the bioassay, FDN were utilized by microbial communities collected from both CH and BW, with complete drawdown of fish-derived NH 4 + in the CH treatment in conjunction with a 3-fold increase in Chl a . FDN provided the necessary nutrients to alleviate P limitation in the BW treatment, resulting in a 13-fold increase in Chl a . Results from this experiment indicate FDN are important N and P sources to the microbial community along the eastern GoM and have important implications for maintenance of associated ichthyotoxic K. brevis blooms.

The West Florida Shelf (WFS) encompasses a range of environments from inshore estuarine to offshore oligotrophic waters, which are frequently the site of large and persistent blooms of the toxic dinoflagellate, Karenia brevis. The goals of this study were to characterize the nitrogen (N) nutrition of plankton across the range of environmental conditions on the WFS, to quantify the percentage of the plankton N demand met through in situ N regeneration, and to determine whether planktonic N nutrition changes when high concentrations of Karenia are present. In the fall of 2007, 2008, and 2009 we measured ambient nutrient concentrations and used stable isotope techniques to measure rates of primary production and uptake rates of inorganic N (ammonium, NH4+, and nitrate, NO3−), and organic N and carbon (C; urea and amino acids, AA) in estuarine, coastal, and offshore waters, as well as coastal sites with Karenia blooms present. In parallel, we also measured rates of in situ N regeneration – NH4+ regeneration, nitrification, and photoproduction of NH4+, nitrite and AA. Based on microscope observations, ancillary measurements, and previous monitoring history, Karenia blooms sampled represented three bloom stages – initiation in 2008, maintenance in 2007, and late maintenance/stationary phase in 2009. Nutrient concentrations were highest at estuarine sampling sites and lowest at offshore sites. Uptake of NH4+ and NO3− provided the largest contribution to N nutrition at all sites. At the non-Karenia sites, in situ rates of NH4+ regeneration and nitrification were generally sufficient to supply these substrates equal to the rates at which they were taken up. At Karenia sites, NO3− was the most important N substrate during the initiation phase, while NH4+ was the most important N form used during bloom maintenance and stationary phases. Rates of NH4+ regeneration were high but insufficient (85 ± 36% of uptake) to support the measured NH4+ uptake at all the Karenia sites although nitrification rates far exceeded uptake rates of NO3−. Taken together our results support the “no smoking gun” nutrient hypothesis that there is no single nutrient source or strategy that can explain Karenia's frequent dominance in the waters where it occurs. Consistent with other papers in this volume, our results indicate that Karenia can utilize an array of inorganic and organic N forms from a number of N sources.

This study represents the most comprehensive assessment of kinetic parameters for Karenia brevis to date as it encompasses natural populations sampled during three different bloom years in addition to cultured strains under controlled conditions. Nitrogen (N) uptake kinetics for ammonium (NH 4 + ), nitrate (NO 3 − ), urea, an amino acid mixture, individual amino acids (glutamate and alanine), and humic substrates were examined for the toxic red tide dinoflagellate, K. brevis , during short term incubations (0.5–1 h) using 15 N tracer techniques. Experiments were conducted using natural populations collected during extensive blooms along the West Florida Shelf in October 2001, 2002, and 2007, and in cultured strains (CCFWC 251 and CCFWC 267) obtained from the Florida Fish and Wildlife Institute culture collection. Kinetic parameters for the maximum uptake velocity ( V max ), half-saturation concentration ( K s ), and the affinity constant ( α ) were determined. The affinity constant is considered a more accurate indicator of substrate affinity at low concentrations. K. brevis took up all organic substrates tested, including N derived from humic substances. Uptake rates of the amino acid mixture and some NO 3 − incubations did not saturate even at the highest substrate additions (50–200 μmol N L −1 ). Based upon the calculated α values, the greatest substrate preference was for NH 4 + followed by NO 3 − ≥ urea, humic compounds and amino acids. The ability of K. brevis to utilize a variety of inorganic and organic substrates likely helps it flourish under a wide range of nutrient conditions from bloom initiation in oligotrophic waters offshore to bloom maintenance near shore where ambient nutrient concentrations may be orders of magnitude greater.

Blooms of Karenia brevis plague the West Florida Shelf (WFS) region in the Gulf of Mexico (GOM) where they exert harmful effects on aquatic biota and humans. Because productivity on the WFS is N limited, new N inputs into the region are thought to trigger blooms of K. brevis. Here we examine the potential for new N inputs via N2 fixation by Trichodesmium and other diazotrophic plankton to contribute to the N demand of K. brevis. Because of possible methodological biases, we also compared N2 fixation rates by cultured Trichodesmium using the 15N2 bubble addition method and the 15N2 saturated seawater. Both methods yielded identical results in 12 and 24 h incubations; however, there was more variability in rate estimates made using the bubble addition method. Pelagic N2 fixation rates by other planktonic diazotrophs ranged from 0 to 13.6 nmol N L−1 d−1, comparable to or higher than rates observed in oligotrophic gyres. These rates should be considered conservative estimates because they were made using the bubble addition method. Integrating over our study area, we estimate that new inputs of N to the WFS via N2 fixation are on the order of 0.011 Tmol N annually. Further, we measured directly the trophic transfer of recently fixed N2 to co-occurring plankton that included K. brevis and found that up to 47% of N2 fixed was transferred to non-diazotrophic plankton even in short (

The toxic HAB dinoflagellate Karenia brevis (Davis) G. Hansen & O. Moestrup (formerly Gymnodinium breve) exhibits a migratory pattern atypical of dinoflagellates: cells concentrate in a narrow (∼0–5 cm) band at the water surface during daylight hours due to phototactic and negative geotactic responses, then disperse downward at night via non-tactic, random swimming. The hypothesis that this daylight surface aggregation behavior significantly influences bacterial and algal productivity and nutrient cycling within blooms was tested during a large, high biomass (chlorophyll a >19 μg L−1) K. brevis bloom in October of 2001 by examining the effects of this surface layer aggregation on inorganic and organic nutrient concentrations, cellular nitrogen uptake, primary and bacterial productivity and the stable isotopic signature (δ15N, δ13C) of particulate material. During daylight hours, concentrations of K. brevis and chlorophyll a in the 0–5 cm surface layer were enhanced by 131% (±241%) and 32.1% (±86.1%) respectively compared with an integrated water sample collection over a 0–1 m depth. Inorganic (NH4, NO3+2, PO4, SiO4) and organic (DOP, DON) nutrient concentrations were also elevated within the surface layer as was both bacterial and primary productivity. Uptake of nitrogen (NH4+, NO3−, urea, dissolved primary amines, glutamine and alanine) compounds by K. brevis was greatest in the surface layer for all compounds tested, with the greatest enhancement evident in urea uptake rates, from 0.08 × 10−5 ng N K. brevis cell−1 h−1 to 3.1 × 10−5 ng N K. brevis cell−1 h−1. These data suggests that this surface aggregation layer is not only an area of concentrated cells within K. brevis blooms, but also an area of increased biological activity and nutrient cycling, especially of nitrogen. Additionally, the classic dinoflagellate migration paradigm of a downward migration for access to elevated NO3− concentrations during the dark period may not apply to certain dinoflagellates such as K. brevis in oligotrophic nearshore areas with no significant nitricline. For these dinoflagellates, concentration within a narrow surface layer in blooms during daylight hours may enhance nutrient supply through biological cycling and photochemical nutrient regeneration.

[1] Dinitrogen (N2) fixation and dissolved organic nitrogen (DON) release rates were measured on fractionated samples (>10 µm and 10 µm and 10 µm and

Dissolved organic nitrogen (DON) produced by the nitrogen-fixer Trichodesmium sp. has the potential to serve as a nitrogen source for the red tide dinoflagellate Karenia brevis. Dis- solved organic matter (DOM) from laboratory cultures of Trichodesmium sp. was isolated, concen- trated and then supplied as a nutrient source to K. brevis cells collected from the Gulf of Mexico. K. brevis abundance increased immediately after Trichodesmium sp. cellular exudate (TCE) addition, allowing the population to double within the first 24 h. There was rapid and complete utilization of the TCE DON as well as ~89% of the TCE dissolved organic phosphorus (DOP). Additionally, ter- minal restriction fragment length polymorphism (TRFLP) was used to assess the bacterial commu- nity response to the addition of TCE . The number of bacterial operational taxonomic units (OTUs) initially increased after the TCE DOM addition, but decreased as K. brevis reached its maximum abundance. Electrospray ionization mass spectrometry (ESI-MS) and Fourier transform ion cy- clotron resonance mass spectrometry (FT-ICR MS) were used to chemically characterize the DOM. Approximately 25% of compounds disappeared within the first 24 h, corresponding to the greatest increase in K. brevis abundance. Using FT-ICR MS, 391 DON and 219 DOP potentially bioavailable compounds were characterized. The bioavailable DON compounds were highly re- duced and 44% had molar ratios indicative of lipid or protein-like compounds. The changes in DON concentration and compound composition show that Tricho desmium sp. provides a sufficient source of nitrogen to directly or indirectly support K. brevis blooms.

Three anthropogenic sources (urban, soil, and industrial runoff) were characterized for bulk nutrient and dissolved organic matter (DOM) composition using wet chemistry and Fourier transform ion cyclotron resonance mass spectrometry. Each source was unique based on its chemical composition. Dissolved inorganic nitrogen (N) comprised 91, 60, and 20 % of the total N pool in the soil, urban, and industrial sources, respectively. The DOM composition was dominated by terrestrial compounds in the soil, condensed hydrocarbons, lipids and proteins in the urban, and lipid-like compounds in the industrial source. A York River (VA) phytoplankton assemblage, dominated by Cochlodinium polykrikoides, was amended with the sources during a 7-day bioassay. There was a doubling of chlorophyll a and/or cell concentrations within 2 days, in the +Urban and +Soil treatments. The + Industrial treatment supported algal growth, but increases in cell abundances were only statistically significant at the end of the experiment (days 5–7), suggesting that this material was less labile to the original York River community than the other anthropogenic nutrient sources, on the relatively short timescale of the study.

Rationale Dissolved organic nitrogen (DON) represents a significant fraction of the total dissolved nitrogen pool in most surface waters and serves as an important nitrogen source for phytoplankton and bacteria. As with other natural organic matter mixtures, ultrahigh resolution Fourier transform ion cyclotron resonance mass spectrometry (FTICRMS) is the only technique currently able to provide molecular composition information on DON. Although electrospray ionization (ESI) is the most commonly used ionization method, it is not very efficient at ionizing most DON components. Methods Positive- and negative-mode atmospheric pressure photoionization (APPI) coupled with ultrahigh resolution FTICRMS at 9.4 T were compared for determining the composition of DON before and after bioassays. Toluene was added as the APPI dopant to the solid-phase DON extracts, producing a final sample that was 90% methanol and 10% toluene by volume. Results Positive-mode (+) APPI proved significantly more efficient at ionizing DON; 62% of the formulas that could be assigned in the positive-ion spectrum contained at least one nitrogen atom vs. 31% in the negative-ion spectrum. FTICR mass spectral data indicated that most of the refractory DON compounds (i.e. nonreactive during the 5 days of the incubation) had molecular compositions representative of lignin-like molecules, while lipid-like and protein-like molecules comprised most of the small reactive component of the DON pool. Conclusions From these data we conclude that (+) APPI FTICRMS is a promising technique for describing the molecular composition of DON mixtures. The technique is particularly valuable in assessing the bioavailability of individual components of DON when combined with bioassays.

In advanced wastewater treatment plants that achieve high levels of nitrogen (N) removal, up to one-third of the N in effluent is organic, herein referred to as effluent organic N (EON). While we know that inorganic N is highly labile, it is unclear what fraction of EON is bioavailable. In this study, we demonstrate the utility of a method that can be used to examine the reactivity of EON in natural receiving waters to better understand both the ecosystem response and the potential bioavailability of EON. The technique is suitable for analyzing polar organic matter in natural waters; electrospray ionization coupled with Fourier transform mass spectrometry. Bioassays were performed on samples collected at the end of the biological process from two wastewater treatment plants achieving advanced N removal. The samples were concentrated, and then added to natural water samples collected from the oligohaline James River, a major tributary of the Chesapeake Bay. Our results demonstrate that while the lignin-like fraction of the effluent dissolved organic matter (some of which contains N) was conserved, a large portion of aliphatic and aromatic compounds containing N was removed (79–100%) during incubations, while other compounds were produced. Furthermore, the two effluents exhibited differences in the degree of degradation and type of degradation, which can be related both to the various processes employed in the two WWTPs and the dramatic differences in the type of influent they received. These findings suggest that EON is highly reactive in the natural environment and that simple assays examining net consumption or production of bulk dissolved organic N pools are inadequate for assessing the bioavailability of EON.

This research addresses the hypothesis that environmental conditions affect temporal connectivity between daily transcription of ribulose-1,5 bisphosphate carboxylase/oxygenase (Rubisco) genes and photosynthetic capacity among phytoplankton. From surface samples collected in the Mississippi River plume (MRP) and Orinoco River plume (ORP), we made size-fractionated measurements of Rubisco (rbcL) mRNA from four phytoplankton groups (heterokonts, haptophytes, Synechococcus, and Prochlorococcus) and chlorophyll-normalized photosynthesis–irradiance parameters (including light-saturated photosynthetic rate—PmaxB), plus nutrient uptake and inorganic carbon. Chlorophyll, photosynthesis, nutrient uptake, and total rbcL mRNA levels were substantially greater in the MRP. Rubisco mRNA and PmaxB exhibited characteristic diel patterns. Regressions with temporally offset data revealed photosynthesis cycles correlated to rbcL mRNA, but with a time lag. This delay was greater in the MRP, and greater among cells >2 μm. At both sites, PmaxB of

Crustacean zooplankton produce dissolved organic matter (DOM) and inorganic nutrients via sloppy feeding, excretion, and fecal pellet leaching. These different mechanisms of the release of metabolic products, however, have never been individually isolated. Our study was designed to determine the relative importance of these different modes on release of dissolved organic carbon (DOC), ammonium (NH4+), and urea from Acartia tonsa calanoid copepods feeding on the diatom Thalassiosira weissflogii. Excretion and sloppy feeding were the dominant modes of DOC production (80 and 20% of total DOC release, respectively) and NH4+ release (93 and 7% of total NH4+ release, respectively). Urea, however, was predominately produced via sloppy feeding and fecal pellet leaching (25% and 62% of total urea release, respectively). Urea contributed 20% of total measured nitrogen (TMN; NH4+ + urea) released from copepods, and constituted 100% of TMN released via fecal pellet leaching, 47% of TMN released via sloppy feeding, and only 3.5% of TMN released via excretion. TMN release was > 100% of copepod body N d− 1, resulting in low DOC:TMN release ratios (4.1 for sloppy feeding, 2.1 for cumulative release of sloppy feeding, excretion, and fecal pellet leaching). Our results suggest that the mechanism of release plays an important role in the amount of different forms of DOM, NH4+, and urea available to bacteria and phytoplankton.

Jellyfish blooms occur in many estuarine and coastal regions and may be increasing in their magnitude and extent worldwide. Voracious jellyfish predation impacts food webs by converting large quantities of carbon (C), fixed by primary producers and consumed by secondary producers, into gelatinous biomass, which restricts C transfer to higher trophic levels because jellyfish are not readily consumed by other predators. In addition, jellyfish release colloidal and dissolved organic matter (jelly-DOM), and could further influence the functioning of coastal systems by altering microbial nutrient and DOM pathways, yet the links between jellyfish and bacterioplankton metabolism and community structure are unknown. Here we report that jellyfish released substantial quantities of extremely labile C-rich DOM, relative to nitrogen (25.6 ± 31.6 C:1N), which was quickly metabolized by bacterioplankton at uptake rates two to six times that of bulk DOM pools. When jelly-DOM was consumed it was shunted toward bacterial respiration rather than production, significantly reducing bacterial growth efficiencies by 10% to 15%. Jelly-DOM also favored the rapid growth and dominance of specific bacterial phylogenetic groups (primarily γ- proteobacteria ) that were rare in ambient waters, implying that jelly-DOM was channeled through a small component of the in situ microbial assemblage and thus induced large changes in community composition. Our findings suggest major shifts in microbial structure and function associated with jellyfish blooms, and a large detour of C toward bacterial CO 2 production and away from higher trophic levels. These results further suggest fundamental transformations in the biogeochemical functioning and biological structure of food webs associated with jellyfish blooms.

Harmful algal blooms (HABs), including toxic species, have been increasing in frequency, range, and duration over the past several decades. The effect of a harmful or toxic algal diet on zooplankton nutrient regeneration, however, has not been previously examined. In this study, we determined the effects of non-bloom and bloom concentrations of non-toxic and toxic cultures of HAB species Prorocentrum minimum and Karlodinium veneficum on grazing and production of dissolved organic carbon (DOC), nitrogen (DON), and phosphorus (DOP), and inorganic nutrients, ammonium (NH4+) and phosphate (PO43−), by the copepod Acartia tonsa and the heterotrophic dinoflagellate Oxyrrhis marina. Ingestion rates of grazers were significantly higher while feeding on bloom algal concentrations compared to non-bloom algal concentrations, but were always below 1% body C d−1 for A. tonsa (ingestion rate range of 0.5–31 ng C individual−1 d−1) and below 2% body C d−1 for O. marina (range of 0.1–8.8 pg C individual−1 d−1). However, rates of inorganic nutrient and dissolved organic matter (DOM) release, when detected, were always >100% of carbon (C), nitrogen (N), and phosphorus (P) ingested. Additionally, the quantity and forms (organic vs. inorganic) of nutrients released by zooplankton were significantly different between non-toxic and toxic algal treatments, and typically higher grazer DOM release occurred while feeding on toxic algal cultures. DOM was the only detected form of nutrient released from O. marina, and DON and DOP were significant portions of total dissolved N and P released for A. tonsa feeding on toxic K. veneficum (69–84% and 73%, respectively). Low grazing rates on all algal diets used in our study, regardless of cell concentration or toxicity, likely resulted in starvation and subsequent catabolism of grazer body tissue. The potential for additional factors affecting variable grazer nutrient release between toxic and non-toxic algal treatments, including algal nutrient quality and direct toxic effects, are discussed. Our results suggest these grazers may not be capable of controlling bloom formation of these HAB species, and that nutrient cycling dynamics in the coastal ocean may change with increases in the presence of harmful and toxic algal blooms.

An important goal of marine biogeochemists is to quantify the rates at which elements cycle through the ocean's diverse microbial assemblage, as well as to determine how these rates vary in time and space. The traditional view that phytoplankton are producers and bacteria are consumers has been found to be overly simplistic, and environmental metagenomics is discovering new and important microbial metabolisms at an accelerating rate. Many nutritional strategies previously attributed to one microorganism or functional group are also or instead carried out by other groups. To tease apart which organism is doing what will require new analytical approaches. Flow cytometry, when combined with other techniques, has great potential for expanding our understanding of microbial interactions because groups can be distinguished optically, sorted, and then collected for subsequent analyses. Herein, we review the advances in our understanding of marine biogeochemistry that have arisen from the use of flow cytometry.

Uptake of inorganic and organic nitrogen (N) by phytoplankton and bacteria was inves- tigated during a mesocosm study conducted in Raunefjord, Norway in April 2005. One mesocosm was fertilized with nitrate and phosphate at a ratio of 16:1 and maintained in the light, while one un- amended light mesocosm served as a control. Dissolved nutrients, phytoplankton and bacterial bio- mass, and phytoplankton community composition were monitored throughout the 26 d experiment. Uptake of 15 N-labeled ammonium and nitrate, and dual-labeled ( 15 N and 13 C) urea and dissolved free amino acids (DFAA) was measured for phytoplankton and bacteria using 2 methods: size fractiona- tion into 0.2-0.8 and >0.8 μm size classes, and flow cytometric sorting based on chlorophyll autoflu- orescence. Prior to fertilization, dissolved inorganic N concentrations were low and comprised ~5% of total dissolved N. Added nitrate was completely utilized in the amended mesocosm within 10 d, stimulating a large bloom of colonial Phaeocystis pouchetii. Ammonium contributed over half of total measured N uptake by phytoplankton and bacteria in both enclosures, while nitrate and urea each supplied roughly 10 to 25%. Overall, DFAA were a negligible N source to phytoplankton but con- tributed 11% to total bacterial N uptake. Bacterial uptake represented a significant portion of total uptake of all N forms, especially urea and DFAA. Comparison of the 2 methods for measuring phyto- plankton versus bacterial uptake demonstrates how the use of 0.8 μm filters can lead to significant overestimation of phytoplankton N uptake due to the retention of bacterial biomass.

Little is known about the relative importance of inorganic and organic nitrogen (N) sources in fueling production of phytoplankton versus heterotrophic bacteria on the continental shelf. This issue was addressed during two diel experiments conducted in the Mid-Atlantic Bight at the Long-term Ecosystem Observatory, LEO-15, off southern New Jersey. Uptake of 15 N-labeled ammonium (NH 4 + ), nitrate (NO 3 − ), and nitrite (NO 2 − ), and dual-labeled ( 15 N and 13 C) urea and dissolved free amino acids was measured in water taken from the surface and bottom mixed layers approximately every 4 h over two 24 h periods in July 2002. Two methods were used to quantify 15 N uptake rates: (1) traditional filtration into various phytoplankton and bacterial size classes, and (2) flow cytometric (FCM) sorting of autotrophic cells based on the presence of chlorophyll autofluorescence. Due to a strong pycnocline, the nutrient composition was quite distinct between the surface and bottom mixed layers. Dissolved organic N (DON) comprised >99% of the total dissolved N (TDN) pool in surface waters, whereas the bottom-water TDN pool was roughly divided between NH 4 + , NO 3 − , and DON. Urea was the dominant N form used by all fractions at the surface, and although phytoplankton >3 μm was responsible for most of the urea uptake, bacterial use was detected using stable isotopes and also suggested by ure C sequence analysis. The majority of ure C sequences recovered from the 0.2–0.8 μm fraction belonged to members of the Alphaproteobacteria (46%), whereas those of the 0.8–3.0 μm size class consisted primarily of Cyanobacteria (70%). In contrast to the surface, N uptake in the bottom layer was dominated by NH 4 + . The bacterial fraction was responsible for 20–49% of the size-fractionated NH 4 + and NO 3 − uptake in surface samples and 36–93% at the bottom. These results suggest that organic N, such as urea, is a viable source of N nutrition to phytoplankton forced to compete with heterotrophic bacteria for limited inorganic N.

Total maximum daily loads for nitrogen (N) are currently being established for the Chesapeake Bay watershed. While we know inorganic N is bioavailable in the environment and therefore its input contributes to cultural eutrophication, the bioavailability of organic N is unclear. Using bioassay experiments, we examined the impact of effluent-derived organic nitrogen (EON) from wastewater treatment plants on natural water samples collected along an estuarine/salinity gradient within the lower Chesapeake Bay watershed. All of the inorganic N and between 31% and 96% of the EON was removed during biotic bioassays within the first 2 days. Further, there was substantial abiotic reactivity of effluent N when it was added to natural water samples. Results demonstrate that organic and inorganic N in effluent is removed to support the growth of microbial communities. These are the first results aimed at assessing the reactivity of EON in natural waters along an estuarine/salinity gradient.

Two different approaches to measuring phytoplankton nitrogen (N) use were compared in late summer 2004 along the main axis of Chesapeake Bay. Uptake of 15N-labeled ammonium and nitrate and dual-labeled (15N and 13C) urea and dissolved free amino acids (DFAA) were measured in surface water samples from upper, mid, and lower bay stations. Two distinct methods were used to assess the relative uptake of N substrates by phytoplankton and correct for bacterial artifacts: (1) traditional filtration using Whatman glass fiber (GF/F) filters and (2) flow cytometric (FCM) sorting of chlorophyll-containing cells. The concentration of dissolved inorganic N (DIN) decreased with distance south along the bay, whereas dissolved organic N (DON) concentrations were relatively constant. Absolute N uptake rates measured using the traditional approach exceeded those of FCM-sorted phytoplankton, thereby suggesting the possibility of bacterial “contamination.” Ammonium was the dominant N form used throughout the transect, although FCM-sorted phytoplankton relied more on urea and DFAA as the ratio of DON/DIN increased toward the bay mouth. Overall, ammonium comprised 74 ± 17%, urea 10 ± 9%, DFAA 9 ± 7%, and nitrate 7 ± 12% of total measured N uptake by phytoplankton. Results suggest that bacteria relied primarily on DFAA and ammonium for N nutrition but also used N from urea at a rate similar to that of phytoplankton, whereas bacterial nitrate uptake was insignificant. On average, phytoplankton uptake of ammonium, urea, and DFAA was overestimated by 61%, 53%, and 135%, respectively, as a result of bacterial retention on GF/F filters.

Large "blooms" of ctenophores (Mnemiopsis leidyi) and scyphomedusae (Chrysaora quinquecirrha) occur throughout the York River, a sub-estuary of Chesapeake Bay. These gelatinous zooplankton blooms can influence carbon (C) and nutrient cycling through excretion of dissolved organic matter (DOM), and inorganic nitrogen (N) and phosphorus (P). We measured dissolved organic carbon, nitrogen and phosphorus (DOC, DON and DOP), ammonium (NH + 4 ) and phosphate (PO 3- 4 ) released by M. leidyi and C. quinquecirrha in the laboratory and estimated their contribution to in situ DOC and inorganic pools. Both species released high amounts of DOC compared with DON and DOP DOM released by Mnemiopsis was C-rich with higher DOC:DON (29:1) compared with the Redfield ratio (6.6C:1N). Daily turnover of DOC and DON in ctenophores was high (25.2% of body C and 18.3% of body N), likely due to mucus production. In contrast, individual Chrysaora released DOC and DON similar to Redfield stoichiometry, but daily turnover of these compounds was low (

The marine nitrogen (N) cycle is a complex network of biological transformations in different N pools. The linkages among these different reservoirs are often poorly understood. Traditional methods for measuring N uptake rely on bulk community properties and cannot provide taxonomic information. 15 N-based stable isotope probing (SIP), however, is a technique that allows detection of uptake of individual N sources by specific microorganisms. In this study we used 15 N SIP methodology to assess the use of different nitrogen substrates by Synechococcus spp. and diatoms on the west Florida shelf. Seawater was incubated in the presence of 15 N-labeled ammonium, nitrate, urea, glutamic acid, and a mixture of 16 amino acids. DNA was extracted and fractionated using CsCl density gradient centrifugation. Quantitative PCR was used to quantify the amounts of Synechococcus and diatom DNA as a function of density, and 15 N tracer techniques were used to measure rates of N uptake by the microbial community. The ammonium, nitrate, urea, and dissolved primary amine uptake rates were 0.077, 0.065, 0.013, and 0.055 μmol N liter −1 h −1 , respectively. SIP data indicated that diatoms and Synechococcus spp. actively incorporated N from [ 15 N]nitrate, [ 15 N]ammonium, and [ 15 N]urea. Synechococcus also incorporated nitrogen from [ 15 N]glutamate and 15 N-amino acids, but no evidence indicating uptake of labeled amino acids by diatoms was detected. These data suggest that N flow in communities containing Synechococcus spp. and diatoms has more plasticity than the new-versus-recycled production paradigm suggests and that these phytoplankters should not be viewed strictly as recycled and new producers, respectively.

Acartia tonsa copepods are not limited to herbivory and can derive up to half their daily ration from predation on heterotrophic ciliates and dinoflagellates. The effects of an omnivorous diet on nutrient regeneration, however, remain unknown. In this study, we fed A. tonsa an exclusively carnivorous diet of either (1a) heterotrophic dinoflagellate Oxyrrhis marina or (1b) Gyrodinium dom- inans, (2) an exclusively herbivorous diet of Thalassiosira weissflogii diatoms, or (3) a mixed omnivo- rous diet. We measured the release rate, composition, and stoichiometry of dissolved organic carbon (DOC), dissolved organic phosphorus (DOP), and nitrogen (urea) in addition to the inorganic nutri- ents, ammonium (NH4 + ) and phosphate (PO4 3- ). Despite similar ingestion rates among treatments, as well as similar C:N ratios of food items, A. tonsa release rates of DOC and NH4 + were highest while feeding on a carnivorous diet and lowest while feeding omnivorously. In contrast, urea, on average, was a higher portion of total nitrogen released in the mixed diet treatment (32 to 59%). DOP release rates were only detectable in diets containing microzooplankton prey. Our results suggest that cope- pod diet plays an important role in determining the quantity and composition of regenerated C, N, and P available to phytoplankton and bacteria. Additionally, the uncoupling of ingestion and nutrient release rates and the variability in released ratios of dissolved C:N:P in our study suggests that stoi- chiometric models based exclusively on predator and prey C:N and N:P ratios may not be adequate in determining stoichiometry of total nutrient release.

The Mattaponi River is part of the York River estuary in Chesapeake Bay. Our objective was to identify the organic matter (OM) sources fueling the lower food web in the tidal freshwater and oligohaline portions of the Mattaponi using the stable isotopes of carbon (C) and nitrogen (N). Over 3 years (2002–2004), we measured zooplankton densities and C and N stable isotope ratios during the spring zooplankton bloom. The river was characterized by a May–June zooplankton bloom numerically dominated by the calanoid copepod Eurytemora affinis and cladocera Bosmina freyi. Cluster analysis of the stable isotope data identified four distinct signatures within the lower food web: freshwater riverine, brackish water, benthic, and terrestrial. The stable isotope signatures of pelagic zooplankton, including E. affinis and B. freyi, were consistent with reliance on a mix of autochthonous and allochthonous OM, including OM derived from vascular plants and humic-rich sediments, whereas macroinvertebrates consistently utilized allochthonous OM. Based on a dual-isotope mixing model, reliance on autochthonous OM by pelagic zooplankton ranged from 20% to 95% of production, declining exponentially with increasing river discharge. The results imply that discharge plays an important role in regulating the energy sources utilized by pelagic zooplankton in the upper estuary. We hypothesize that this is so because during high discharge, particulate organic C loading to the upper estuary increased and phytoplankton biomass decreased, thereby decreasing phytoplankton availability to the food web.

A mesocosm experiment was conducted during the spring of 2003 in Raunefjord, west Norway. Inorganic nutrients (16 µmol l -1 nitrate, 1 µmol l -1 phosphate) were added to 2 enclosures with 10% of those concentrations (1.6 µmol l -1 nitrate, 0.1 µmol l -1 phosphate) added daily thereafter; a third unamended mesocosm was used as a control. Nitrogen (N) (ammonium, nitrate, urea and amino acid) uptake rates for >0.8 µm (largely composed of phytoplankton) and 0.2 to 0.8 µm (largely composed of bacteria) size classes were measured, as well as nutrient, chlorophyll, phytoplankton and microzooplankton concentrations. The nutrient additions initiated a process resulting in a large bloom of Phaeocystis pouchetii colonies. There was a 2.5 wk lag, during which a smaller bloom of phototrophic flagellates, followed by diatoms, formed in all mesocosms; diatoms increased until sili- cic acid was depleted. After the flagellate and diatom bloom dissipated, the mesocosms were depleted of inorganic N. Dissolved organic N (DON) remained constant in all mesocosms during the flagellate and diatom bloom but increased with the onset of the large P. pouchetii bloom, largely due to release of urea and amino acids, which made up >93% of the DON pool at the end of the experi- ment. Uptake rates of urea often accounted for the largest percentage of total N uptake in all meso- cosms for both the >0.8 µm and 0.2 to 0.8 µm size classes, generally providing the largest percentage (up to 88%) of bacterial N demand of any substrate. This was particularly true during the Phaeocys- tis bloom when urea uptake rates were up to 65 times greater than the other N substrates. We observed a switch from new production, when nitrate uptake dominated during the diatom bloom, to regenerated production during the Phaeocystis bloom. The present study adds to the growing body of evidence that organic N is a substantial source of N for both phytoplankton and bacteria.

Our objective was to quantify the contribution of autochthonous, locally-produced phytoplankton, and allochthonous, terrestrial-derived organic matter (OM) to the production of young-of-year (YOY) American shad(Alosa sapidissima) using stable isotopes. We measured the carbon and nitrogen stable isotope composition of YOY American shad in the tidal fresh water of the Mattaponi River, a tributary in the York River estuary, during three consecutive years. The isotopic ratios of larval American shad varied among years, indicating a switch from reliance on a primarily autochthonous food web pathway during low and moderate discharge years (50–90%; 2002, 2004) to a primarily allochthonous pathway during a high discharge year (< 35% phytoplankton; 2003). Reliance on phytoplankton by larval fish declined exponentially with increasing Mattaponi River discharge. In 2003, juvenile production was also supported by allochthonous OM, though autochthonous phytoplankton accounted for an increasingly large fraction during June through August, up to 40–55%. We also found a long-term, positive relationship between the duration of above average flow during April through June in the Mattaponi River and a corresponding index of juvenile American shad abundance. The largest American shad cohort recorded since 1967 was observed in 2003, a high discharge year. The production of this cohort was largely supported by allochthonous OM. The results suggest an important link between river discharge, energy flow, and recruitment, wherein high discharge favors reliance on terrestrial carbon by YOY American shad, owing to changes in zooplankton diet, macroinvertebrate abundance, or both, and also favors high American shad abundance.

Bacterioplankton nutrient metabolism in the Eastern Tropical North Pacific (ETNP) was assessed using specific activities of intracellular nitrogen (N) assimilation enzymes and hydrolytic ectoenzymes during amendment experiments, mesocosms, and diel studies of in situ rates. Glutamine synthetase (GS) and assimilatory nitrate reductase (ANR) were used to investigate N bioavailability, alkaline phosphatase (AP) to assess phosphorous (P) bioavailability and β-glucosidase (β-Glu) to detect shifts in the use of labile dissolved organic carbon (DOC). Conditions regulating activity of each enzyme were tested using incubations of

River plumes deliver large quantities of nutrients to oligotrophic oceans, often resulting in significant CO(2) drawdown. To determine the relationship between expression of the major gene in carbon fixation (large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase, RuBisCO) and CO(2) dynamics, we evaluated rbcL mRNA abundance using novel quantitative PCR assays, phytoplankton cell analyses, photophysiological parameters, and pCO(2) in and around the Mississippi River plume (MRP) in the Gulf of Mexico. Lower salinity (30-32) stations were dominated by rbcL mRNA concentrations from heterokonts, such as diatoms and pelagophytes, which were at least an order of magnitude greater than haptophytes, alpha-Synechococcus or high-light Prochlorococcus. However, rbcL transcript abundances were similar among these groups at oligotrophic stations (salinity 34-36). Diatom cell counts and heterokont rbcL RNA showed a strong negative correlation to seawater pCO(2). While Prochlorococcus cells did not exhibit a large difference between low and high pCO(2) water, Prochlorococcus rbcL RNA concentrations had a strong positive correlation to pCO(2), suggesting a very low level of RuBisCO RNA transcription among Prochlorococcus in the plume waters, possibly due to their relatively poor carbon concentrating mechanisms (CCMs). These results provide molecular evidence that diatom/pelagophyte productivity is largely responsible for the large CO(2) drawdown occurring in the MRP, based on the co-occurrence of elevated RuBisCO gene transcript concentrations from this group and reduced seawater pCO(2) levels. This may partly be due to efficient CCMs that enable heterokont eukaryotes such as diatoms to continue fixing CO(2) in the face of strong CO(2) drawdown. Our work represents the first attempt to relate in situ microbial gene expression to contemporaneous CO(2) flux measurements in the ocean.