Rare but large bivalves alter benthic respiration and nutrient recycling in riverine sediments

Bioturbation studies have generally analyzed small and abundant organisms while the contribution to the benthic metabolism by rare, large macrofauna has received little attention. We hypothesize that large, sporadic bivalves may represent a hot spot for benthic processes due to a combination of direct and indirect effects as their metabolic and bioturbation activities. Intact riverine sediments with and without individuals of the bivalve Sinanodonta woodiana were collected in a reach with transparent water, where the occurrence of the mollusk was clearly visible. The bivalve metabolism and its effects on sedimentary fluxes of dissolved gas and nutrients were measured via laboratory incubations of intact cores under controlled conditions. S. woodiana contributed significantly to O₂ and TCO₂ benthic fluxes through its respiration and to \({\text{NH}}_{4}^{ + }\), SRP and SiO₂ regeneration via its excretion. The bivalve significantly stimulated also microbial denitrification and determined a large efflux of CH₄, likely due a combination of bioturbation and biodeposition activities or to anaerobic metabolism within the mollusk gut. This study demonstrates that a few, large individuals of this bivalve produce significant effects on aerobic and anaerobic benthic metabolism and nutrient mobilization. Random sediment sampling in turbid waters seldom catches these important effects due to low densities of large fauna.     

Density of Monoporeia affinis and biogeochemistry in Baltic Sea sediments

This study focused on effects from Monoporeia affinis reworking and ventilation activities on benthic fluxes and mineralization processes during a simulated bloom event. The importance of M. affinis density for benthic solute (O₂, ΣNO₂⁻ + NO₃⁻, NH₄⁺ and HPO₄²⁻) fluxes and sediment reactivity (mobilization of NH₄⁺ and HPO₄²⁻) following additions of organic material to the sediment surface was experimentally investigated using sediment-water and closed sediment (jar) incubations. Three different densities of M. affinis were used to resemble a low, medium and high density situation (1300, 2500 and 6400 ind. m^− 2, respectively) of a natural amphipod community. The degradation of phytodetritus (Tetraselmis sp., 5 g C m^− 2) added to the sediment surface was followed over a period of 20 days. Benthic solute fluxes of O₂, ΣNO₂⁻ + NO₃⁻ and NH₄⁺ were generally progressively stimulated with increasing number of M. affinis, while no such correlation was found for HPO₄²⁻. Solute fluxes were initially enhanced 1 to 2 days after the addition of phytodetritius, caused by mineralization of the most labile organic material and a food-stimulated irrigation by the amphipods. There was no effect from the activity of M. affinis on total denitrification (Dtot = Dn + Dw) or denitrification utilizing nitrate from coupled nitrification/denitrification (Dn) for any of the densities examined. Denitrification utilizing overlying water nitrate (Dw) was only about 10% of Dtot. Dw was significantly enhanced for the highest M. affinis density investigated. The reactivity of the sediment decreased progressively with increasing density of M. affinis and with time of the experiment. However, enhanced ammonium production at least 6 days after the organic addition indicated excretion of N-containing organic compounds by M. affinis. In conclusion, large spatial and temporal variations in density of M. affinis may be of significant importance for benthic solute fluxes and overall mineralization of organic material in Baltic Sea sediments.     

Multi-scale measurements and modeling of denitrification in streams with varying flow and nitrate concentration in the upper Mississippi River basin, USA

Denitrification is an important net sink for NO₃ ^? in streams, but direct measurements are limited and in situ controlling factors are not well known. We measured denitrification at multiple scales over a range of flow conditions and NO₃ ^? concentrations in streams draining agricultural land in the upper Mississippi River basin. Comparisons of reach-scale measurements (in-stream mass transport and tracer tests) with local-scale in situ measurements (pore-water profiles, benthic chambers) and laboratory data (sediment core microcosms) gave evidence for heterogeneity in factors affecting benthic denitrification both temporally (e.g., seasonal variation in NO₃ ^? concentrations and loads, flood-related disruption and re-growth of benthic communities and organic deposits) and spatially (e.g., local stream morphology and sediment characteristics). When expressed as vertical denitrification flux per unit area of streambed (U _denit, in μmol N m^?2 h^?1), results of different methods for a given set of conditions commonly were in agreement within a factor of 2–3. At approximately constant temperature (~20 ± 4°C) and with minimal benthic disturbance, our aggregated data indicated an overall positive relation between U _denit (~0–4,000 μmol N m^?2 h^?1) and stream NO₃ ^? concentration (~20–1,100 μmol L^?1) representing seasonal variation from spring high flow (high NO₃ ^?) to late summer low flow (low NO₃ ^?). The temporal dependence of U _denit on NO₃ ^? was less than first-order and could be described about equally well with power-law or saturation equations (e.g., for the unweighted dataset, U _denit ≈26 * [NO₃ ^?]^0.44 or U _denit ≈640 * [NO₃ ^?]/[180 + NO₃ ^?]; for a partially weighted dataset, U _denit ≈14 * [NO₃ ^?]^0.54 or U _denit ≈700 * [NO₃ ^?]/[320 + NO₃ ^?]). Similar parameters were derived from a recent spatial comparison of stream denitrification extending to lower NO₃ ^? concentrations (LINX2), and from the combined dataset from both studies over 3 orders of magnitude in NO₃ ^? concentration. Hypothetical models based on our results illustrate: (1) U _denit was inversely related to denitrification rate constant (k1_denit, in day^?1) and vertical transfer velocity (v _f,denit, in m day^?1) at seasonal and possibly event time scales; (2) although k1_denit was relatively large at low flow (low NO₃ ^?), its impact on annual loads was relatively small because higher concentrations and loads at high flow were not fully compensated by increases in U _denit; and (3) although NO₃ ^? assimilation and denitrification were linked through production of organic reactants, rates of NO₃ ^? loss by these processes may have been partially decoupled by changes in flow and sediment transport. Whereas k1_denit and v _f,denit are linked implicitly with stream depth, NO₃ ^? concentration, and(or) NO₃ ^? load, estimates of U _denit may be related more directly to field factors (including NO₃ ^? concentration) affecting denitrification rates in benthic sediments. Regional regressions and simulations of benthic denitrification in stream networks might be improved by including a non-linear relation between U _denit and stream NO₃ ^? concentration and accounting for temporal variation.     

Coupling Nutrient Uptake and Energy Flow in Headwater Streams

Nutrient cycling and energy flow in ecosystems are tightly linked through the metabolic processes of organisms. Greater uptake of inorganic nutrients is expected to be associated with higher rates of metabolism [gross primary production (GPP) and respiration (R)], due to assimilatory demand of both autotrophs and heterotrophs. However, relationships between uptake and metabolism should vary with the relative contribution of autochthonous and allochthonous sources of organic matter. To investigate the relationship between metabolism and nutrient uptake, we used whole-stream and benthic chamber methods to measure rates of nitrate–nitrogen (NO₃–N) uptake and metabolism in four headwater streams chosen to span a range of light availability and therefore differing rates of GPP and contributions of autochthonous carbon. We coupled whole-stream metabolism with measures of NO₃–N uptake conducted repeatedly over the same stream reach during both day and night, as well as incubating benthic sediments under both light and dark conditions. NO₃–N uptake was generally greater in daylight compared to dark conditions, and although day-night differences in whole-stream uptake were not significant, light–dark differences in benthic chambers were significant at three of the four sites. Estimates of N demand indicated that assimilation by photoautotrophs could account for the majority of NO₃–N uptake at the two sites with relatively open canopies. Contrary to expectations, photoautotrophs contributed substantially to NO₃–N uptake even at the two closed-canopy sites, which had low values of GPP/R and relied heavily on allochthonous carbon to fuel R.     

An integrated network analysis reveals that nitric oxide reductase prevents metabolic cycling of nitric oxide by Pseudomonas aeruginosa

Nitric oxide (NO) is a chemical weapon within the arsenal of immune cells, but is also generated endogenously by different bacteria. Pseudomonas aeruginosa are pathogens that contain an NO-generating nitrite (NO₂⁻) reductase (NirS), and NO has been shown to influence their virulence. Interestingly, P. aeruginosa also contain NO dioxygenase (Fhp) and nitrate (NO₃⁻) reductases, which together with NirS provide the potential for NO to be metabolically cycled (NO→NO₃⁻→NO₂⁻→NO). Deeper understanding of NO metabolism in P. aeruginosa will increase knowledge of its pathogenesis, and computational models have proven to be useful tools for the quantitative dissection of NO biochemical networks. Here we developed such a model for P. aeruginosa and confirmed its predictive accuracy with measurements of NO, O₂, NO₂⁻, and NO₃⁻ in mutant cultures devoid of Fhp or NorCB (NO reductase) activity. Using the model, we assessed whether NO was metabolically cycled in aerobic P. aeruginosa cultures. Calculated fluxes indicated a bottleneck at NO₃⁻, which was relieved upon O₂ depletion. As cell growth depleted dissolved O₂ levels, NO₃⁻ was converted to NO₂⁻ at near-stoichiometric levels, whereas NO₂⁻ consumption did not coincide with NO or NO₃⁻ accumulation. Assimilatory NO₂⁻ reductase (NirBD) or NorCB activity could have prevented NO cycling, and experiments with ΔnirB, ΔnirS, and ΔnorC showed that NorCB was responsible for loss of flux from the cycle. Collectively, this work provides a computational tool to analyze NO metabolism in P. aeruginosa, and establishes that P. aeruginosa use NorCB to prevent metabolic cycling of NO.     

Primary Productivity of Phytoplankton,Environmental Covariates and Fish Growth in West Bengal Ponds under a Polyculture System

Measurements of primary production by phytoplankton in four fish ponds over a period of one year revealed a distinct spatial variation in rate. The ponds were used for polyculture, monoculture, or a traditional system of fish culture. The rates of gross photosynthesis were highly correlated with temperature, free CO₂, oxygen, inorganic nitrogen (NH₄⁺, NO₂⁻, and NO₃⁻) and PO₄-P. It was shown by step-wise multiple regression analysis that in the polyculture pond with a predominantly autotrophic metabolism, gross photosynthesis was the most significant factor correlated with fish growth. In another polyculture system in which the metabolism was mainly hetero-trophic, the influence of the photosynthesis rate was slight.     

The Uptake of NO3?, NO2?, and NH4+ by Intact Wheat (Triticum aestivum) Seedlings : I. Induction and Kinetics of Transport Systems

The inducibility and kinetics of the NO₃⁻, NO₂⁻, and NH₄⁺ transporters in roots of wheat seedlings (Triticum aestivum cv Yercora Rojo) were characterized using precise methods approaching constant analysis of the substrate solutions. A microcomputer-controlled automated high performance liquid chromatography system was used to determine the depletion of each N species (initially at 1 millimolar) from complete nutrient solutions. Uptake rate analyses were performed using computerized curve-fitting techniques. More precise estimates were obtained for the time required for and the extent of the induction of each transporter. Up to 10 and 6 hours, respectively, were required to achieve apparent full induction of the NO₃⁻ and NO₂⁻ transporters. Evidence for substrate inducibility of the NH₄⁺ transporters requiring 5 hours is presented. The transport of NO₃⁻ was mediated by a dual system (or dual phasic), whereas only single systems were found for transport of NO₂⁻ and NH₄⁺. The K_m values for NO₃⁻, NO₂⁻, and NH₄⁺ were, respectively, 0.027, 0.054, and 0.05 millimolar. The K_m for mechanism II of NO₃⁻ transport could not be defined in this study as it exhibited only apparent first order kinetics up to 1 millimolar.     

Benthic decomposition of Zostera marina roots: a controlled laboratory experiment

The initial benthic decomposition of Zostera marina roots was studied in a controlled flow-through chamber experiment for 23 days. Sediment chambers without added roots served as controls. The inflowing and outflowing artificial seawater (ASW) was analyzed for O₂, ΣCO₂, urea-N, NH₄⁺ and NO₂⁻+NO₃⁻. Sediment profiles of Eh, particulate organic carbon (POC) and nitrogen, dissolved organic nitrogen (DON), dissolved free amino acids (DFAA), urea-N, NH₄⁺, DFAA and urea turnover rates, sulfate reduction and counts of total anaerobic heterotrophic bacteria and different functional groups were determined. Fluxes of O₂, ΣCO₂, urea-N and NH₄⁺ were stimulated during root decomposition compared to the unamended control. There were indications of stimulated bacterial growth based on counts of total anaerobic heterotrophic bacteria, anaerobic phosphatase utilizers, ammonifyers and sulfate reducers. Independent estimates of nitrogen and carbon incorporation into bacterial biomass during root decomposition indicate that a major fraction of the nitrogen for microbial growth was mobilized from the indigenous particulate organic nitrogen (PON) pool, whereas the energy source for bacterial growth was mainly obtained from the added eelgrass roots. Most of the nitrogen mineralized during root decomposition was incorporated into the bacterial biomass resulting in a low efflux of urea-N and inorganic nitrogen from the sediment to the water column.     

Immobilization ofPhotobacterium phosphoreum for monitoring of toxic substances

A new sensing system based on the immobilization of luminescent bacteria,Photobacterium phosphoreum, was proposed for continuous real-time monitoring of pollutants. The response curves demonstrate thatPhotobacterium phosphoreum immobilized on the strontium alginate was very sensitive to seven reference chemicals used. The significant inhibitory concentrations for bioluminescence emission were 5 ppm for Pb(NO₃)₂, NiCl₂, CdCl₂, 50 ppm for NaAsO₂, 0.1 ppm for HgCl₂, 0.5 ppm for pentachlorophenol and less than 5 ppm for SDS, respectively. The alginate mixed-cells (AMC) retained their luminescence during experimental period (29 days) under storage condition of ? 80°C. The variables affecting performance of continuous flow through monitoring (CFTM) were optimized in order to ensure stability and efficiency. The flow through cell with strontium-alginate immobilized luminescent bacteria was tested with salicylate and 4-nitrophenol and a rapid response of luminescence was recorded by time drivemode in bioluminescence spectrometer after exposure to both toxicants.     

Ecology of Thioploca spp.: Nitrate and Sulfur Storage in Relation to Chemical Microgradients and Influence of Thioploca spp. on the Sedimentary Nitrogen Cycle

Microsensors, including a recently developed NO₃⁻ biosensor, were applied to measure O₂ and NO₃⁻ profiles in marine sediments from the upwelling area off central Chile and to investigate the influence of Thioploca spp. on the sedimentary nitrogen metabolism. The studies were performed in undisturbed sediment cores incubated in a small laboratory flume to simulate the environmental conditions of low O₂, high NO₃⁻, and bottom water current. On addition of NO₃⁻ and NO₂⁻, Thioploca spp. exhibited positive chemotaxis and stretched out of the sediment into the flume water. In a core densely populated with Thioploca, the penetration depth of NO₃⁻ was only 0.5 mm and a sharp maximum of NO₃⁻ uptake was observed 0.5 mm above the sediment surface. In sediments with only few Thioploca spp., NO₃⁻ was detectable down to a depth of 2 mm and the maximum consumption rates were observed within the sediment. No chemotaxis toward nitrous oxide (N₂O) was observed, which is consistent with the observation that Thioploca does not denitrify but reduces intracellular NO₃⁻ to NH₄⁺. Measurements of the intracellular NO₃⁻ and S⁰ pools in Thioploca filaments from various depths in the sediment gave insights into possible differences in the migration behavior between the different species. Living filaments containing significant amounts of intracellular NO₃⁻ were found to a depth of at least 13 cm, providing final proof for the vertical shuttling of Thioploca spp. and nitrate transport into the sediment.     

Nitrification and Denitrification in Lake and Estuarine Sediments Measured by the 15N Dilution Technique and Isotope Pairing

The transformation of nitrogen compounds in lake and estuarine sediments incubated in the dark was analyzed in a continuous-flowthrough system. The inflowing water contained ¹⁵NO₃^-, and by determination of the isotopic composition of the N₂, NO₃^-, and NH₄⁺ pools in the outflowing water, it was possible to quantify the following reactions: total NO₃^- uptake, denitrification based on NO₃^- from the overlying water, nitrification, coupled nitrification-denitrification, and N mineralization. In sediment cores from both lake and estuarine environments, benthic microphytes assimilated NO₃^- and NH₄⁺ for a period of 25 to 60 h after darkening. Under steady-state conditions in the dark, denitrification of NO₃^- originating from the overlying water accounted for 91 to 171 μmol m^-2 h^-1 in the lake sediments and for 131 to 182 μmol m^-2 h^-1 in the estuarine sediments, corresponding to approximately 100% of the total NO₃^- uptake for both sediments. It seems that high NO₃^- uptake by benthic microphytes in the initial dark period may have been misinterpreted in earlier investigations as dissimilatory reduction to ammonium. The rates of coupled nitrification-denitrification within the sediments contributed to 10% of the total denitrification at steady state in the dark, and total nitrification was only twice as high as the coupled process.     

Oxygen stress in Salvinia natans: Interactive effects of oxygen availability and nitrogen source

The ability of Salvinia natans (L.) All. to tolerate growth in oxic, hypoxic and anoxic nutrient solutions when supplied with either NH₄⁺ or NO₃^? were studied in the laboratory to test the hypothesis that inorganic N-source affects the response of the plants to O₂ deprivation. The relative growth rate (RGR) was significantly reduced in the anoxic treatment, but in the hypoxic treatment RGR was only slightly affected. The NH₄⁺ fed plants generally had a higher shoot to root ratio than the NO₃^? fed plants, and highest in the anoxic treatment. Plants had more roots and larger leaves when supplied with NH₄⁺ as compared with NO₃^?, particularly in the oxic treatment, and root length was most affected by O₂ deprivation for NO₃^? fed plants. Cell walls in the endodermis, the bundle sheath and the cortex adjacent to endodermis developed thickened sclerenchymatous walls when deprived of O₂, and more so in plants supplied with NO₃^?. Plants lost chlorophylls, had lower rates of photosynthetic electron transport (ETR_max) and lower quantum yields (Fv/Fm ratios) when grown in anoxic solutions, and the negative effects were mildest for NO₃^? fed plants suggesting that NO₃^? may be used as an alternative e^?-acceptor in non-cyclic electron transport in the chloroplasts. Overall S. natans grew best on NH₄⁺, but it also grew well on NO₃^?, and the O₂ stress symptoms differed somewhat between NH₄⁺ fed and NO₃^? fed plants. However, because N-form itself significantly influenced morphology and cell metabolism, it was impossible to conclusively identify the role of N-form for the O₂ stress reactions. S. natans is not well-adapted to grow in O₂ deficient waters and will not tolerate completely anoxic conditions as will prevail in waters receiving high loadings of organic pollutants such as livestock wastewater.     

Identification and Activities In Situ of Nitrosospira and Nitrospira spp. as Dominant Populations in a Nitrifying Fluidized Bed Reactor

Bacterial aggregates from a chemolithoautotrophic, nitrifying fluidized bed reactor were investigated with microsensors and rRNA-based molecular techniques. The microprofiles of O₂, NH₄⁺, NO₂⁻, and NO₃⁻ demonstrated the occurrence of complete nitrification in the outer 125 μm of the aggregates. The ammonia oxidizers were identified as members of the Nitrosospira group by fluorescence in situ hybridization (FISH). No ammonia- or nitrite-oxidizing bacteria of the genus Nitrosomonas or Nitrobacter, respectively, could be detected by FISH. To identify the nitrite oxidizers, a 16S ribosomal DNA clone library was constructed and screened by denaturing gradient gel electrophoresis and selected clones were sequenced. The organisms represented by these sequences formed two phylogenetically distinct clusters affiliated with the nitrite oxidizer Nitrospira moscoviensis. 16S rRNA-targeted oligonucleotide probes were designed for in situ detection of these organisms. FISH analysis showed that the dominant populations of Nitrospira spp. and Nitrosospira spp. formed separate, dense clusters which were in contact with each other and occurred throughout the aggregate. A second, smaller, morphologically and genetically different population of Nitrospira spp. was restricted to the outer nitrifying zones.     

Bacterium-Based NO2− Biosensor for Environmental Applications

A sensitive NO₂⁻ biosensor that is based on bacterial reduction of NO₂⁻ to N₂O and subsequent detection of the N₂O by a built-in electrochemical N₂O sensor was developed. Four different denitrifying organisms lacking NO₃⁻ reductase activity were assessed for use in the biosensor. The relevant physiological aspects examined included denitrifying characteristics, growth rate, NO₂⁻ tolerance, and temperature and salinity effects on the growth rate. Two organisms were successfully used in the biosensor. The preferred organism was Stenotrophomonas nitritireducens, which is an organism with a denitrifying pathway deficient in both NO₃⁻ and N₂O reductases. Alternatively Alcaligenes faecalis could be used when acetylene was added to inhibit its N₂O reductase. The macroscale biosensors constructed exhibited a linear NO₂⁻ response at concentrations up to 1 to 2 mM. The detection limit was around 1 μM NO₂⁻, and the 90% response time was 0.5 to 3 min. The sensor signal was specific for NO₂⁻, and interference was observed only with NH₂OH, NO, N₂O, and H₂S. The sensor signal was affected by changes in temperature and salinity, and calibration had to be performed in a system with a temperature and an ionic strength comparable to those of the medium analyzed. A broad range of water bodies could be analyzed with the biosensor, including freshwater systems, marine systems, and oxic-anoxic wastewaters. The NO₂⁻ biosensor was successfully used for long-term online monitoring in wastewater. Microscale versions of the NO₂⁻ biosensor were constructed and used to measure NO₂⁻ profiles in marine sediment.     

Estimation of Nitrification and Denitrification from Microprofiles of Oxygen and Nitrate in Model Sediment Systems

The coupling between nitrification and denitrification and the regulation of these processes by oxygen were studied in freshwater sediment microcosms with O₂ and NO₃^- microsensors. Depth profiles of nitrification (indicated as NO₃^- production), denitrification (indicated as NO₃^- consumption), and O₂ consumption activities within the sediment were calculated from the measured concentration profiles. From the concentration profiles, it was furthermore possible to distinguish between the rate of denitrification based on the diffusional supply of NO₃^- from the overlying water and the rate based on NO₃^- supplied by benthic nitrification (D_w and D_n, respectively). An increase in O₂ concentration caused a deeper O₂ penetration while a decrease in D_w and an increase in D_n were observed. The relative importance for total denitrification of NO₃^- produced by nitrification thus increased compared with NO₃^- supplied from the water phase. The decrease in D_w at high oxygen was due to an increase in diffusion path for NO₃^- from the overlying water to the denitrifying layers in the anoxic sediment. At high O₂ concentrations, nitrifying activity was restricted to the lower part of the oxic zone where there was a continuous diffusional supply of NH₄⁺ from deeper mineralization processes, and the long diffusion path from the nitrification zone to the overlying water compared with the path to the denitrifying layers led to a stimulation in D_n.     

Uptake of dissolved inorganic and organic nitrogen by the benthic toxic dinoflagellate Ostreopsis cf. ovata

Environmental factors that shape dynamics of benthic toxic blooms are largely unknown. In particular, for the toxic dinoflagellate Ostreopsis cf. ovata, the importance of the availability of nutrients and the contribution of the inorganic and organic pools to growth need to be quantified in marine coastal environments. The present study aimed at characterizing N-uptake of dissolved inorganic and organic sources by O. cf. ovata cells, using the ¹⁵N-labelling technique. Experiments were conducted taking into account potential interactions between nutrient uptake systems as well as variations with the diel cycle. Uptake abilities of O. cf. ovata were parameterized for ammonium (NH₄⁺), nitrate (NO₃⁻) and N-urea, from the estimation of kinetic and inhibition parameters. In the range of 0 to 10 μmol N L⁻¹, kinetic curves showed a clear preference pattern following the ranking NH₄⁺ > NO₃⁻ > N-urea, where the preferential uptake of NH₄⁺ relative to NO₃⁻ was accentuated by an inhibitory effect of NH₄⁺ concentration on NO₃⁻ uptake capabilities. Conversely, under high nutrient concentrations, the preference for NH₄⁺ relative to NO₃⁻ was largely reduced, probably because of the existence of a low-affinity high capacity inducible NO₃⁻ uptake system. Ability to take up nutrients in darkness could not be defined as a competitive advantage for O. cf. ovata. Species competitiveness can also be defined from nutrient uptake kinetic parameters. A strong affinity for NH₄⁺ was observed for O. cf. ovata cells that may partly explain the success of this toxic species during the summer season in the Bay of Villefranche-sur-mer (France).