Rapid Assessment of Intertidal Wetland Sediments

Urbanization of coastal areas poses a severe threat to ecologically valuable intertidal wetlands. This paper presents a pragmatic approach called Rapid Assessment for Intertidal Wetland Sediments (RAITWS) for evaluating the sediment quality of intertidal wetlands. RAITWS involves construction of reference groups, selection of a subset of environmental variables, matching of test sites to reference groups, prediction of the benthic fauna community structure (e.g. of macroinvertebrates) at test sites, evaluation of the Observation to Expectation ratio (O/E ratio), quantification of environmental variables with series of dynamic numerical models, and interpretation of the O/E findings. The proposed method extends the existing rapid biological assessment approach from static to dynamic applications. In particular, RAITWS provides a fast method of assessing intertidal wetland sites which are undergoing ecological change due to nearby coastal development.     

Denitrification in San Francisco Bay Intertidal Sediments

The acetylene block technique was employed to study denitrification in intertidal estuarine sediments. Addition of nitrate to sediment slurries stimulated denitrification. During the dry season, sediment-slurry denitrification rates displayed Michaelis-Menten kinetics, and ambient NO₃⁻ + NO₂⁻ concentrations (≤26 μM) were below the apparent K_m (50 μM) for nitrate. During the rainy season, when ambient NO₃⁻ + NO₂⁻ concentrations were higher (37 to 89 μM), an accurate estimate of the K_m could not be obtained. Endogenous denitrification activity was confined to the upper 3 cm of the sediment column. However, the addition of nitrate to deeper sediments demonstrated immediate N₂O production, and potential activity existed at all depths sampled (the deepest was 15 cm). Loss of N₂O in the presence of C₂H₂ was sometimes observed during these short-term sediment incubations. Experiments with sediment slurries and washed cell suspensions of a marine pseudomonad confirmed that this N₂O loss was caused by incomplete blockage of N₂O reductase by C₂H₂ at low nitrate concentrations. Areal estimates of denitrification (in the absence of added nitrate) ranged from 0.8 to 1.2 μmol of N₂ m⁻² h⁻¹ (for undisturbed sediments) to 17 to 280 μmol of N₂ m⁻² h⁻¹ (for shaken sediment slurries).     

Characterization of beta-Glucosidase Activity in Intertidal Marine Sediments

Glycoside derivatives of 4-methylumbelliferone (MUF) were used to characterize the polysaccharidase enzyme systems present in sediments from an intertidal mud flat. The formation of highly fluorescent MUF on hydrolysis of the various glycosides was determined at low substrate concentrations (<1 muM) and with short incubation periods (>5 min). The hydrolysis of MUF-beta-d-glucose in sediments from depth intervals of 0 to 2 cm was insensitive to the presence of oxygen, dissolved sulfide, and iron; magnesium and calcium were stimulatory, however. A pronounced temperature optimum was observed at 40 degrees C, a salinity optimum at 30 per thousand, and a pH optimum at 8.5. Rates of hydrolysis were completely inhibited by the addition of mercuric chloride and sodium azide, but only partially inhibited by toluene and the specific beta-glucosidase inhibitor delta-1,5-gluconolactone. The response to delta-1,5-gluconolactone suggested that about 50% of the observed hydrolysis of MUF-beta-d-glucoside was due to exo- and endoglucanases. A wide variety of hydrolytic activities was observed, with at least some nonspecificity occurring in the case of MUF-beta-d-fucoside. Depth profiles indicated maximal activity in surface sediments with a rapid decline below 2 cm. MUF-glycosides provided a convenlent tool for initial analyses of the dynamics and controls of polymer hydrolysis in marine sediments.     

Spatial and Temporal Variation of Phenanthrene-Degrading Bacteria in Intertidal Sediments

Phenanthrene-degrading bacteria were isolated from a 1-m² intertidal sediment site in Boston Harbor. Samples were taken six times over 2 years. A total of 432 bacteria were isolated and characterized by biochemical testing. When clustered on the basis of phenotypic characteristics, the isolates could be separated into 68 groups at a similarity level of approximately 70%. Several groups (a total of 200 isolates) corresponded to well-characterized species belonging the genera Vibrio and Pseudomonas. Only 51 of the 437 isolates (<11.7% of the total) hybridized to a DNA probe that encodes the upper pathway of naphthalene and phenanthrene degradation in Pseudomonas putida NCIB 9816. A cluster analysis indicated that the species composition of the phenanthrene-degrading community changed significantly from sampling date to sampling date. At one sampling time, 12 6-mm-diameter core subsamples were taken within the 1-m² site to determine the spatial variability of the degrading communities. An analysis of molecular variance, performed with the phenotypic characteristics, indicated that only 6% of the variation occurred among the 12 subsamples, suggesting that the subsamples were almost identical in composition. We concluded that the communities of phenanthrene-degrading bacteria in the sediments are very diverse, that the community structure undergoes significant change with time but does not vary significantly on a spatial scale of centimeters, and that the predominant genes that encode phenanthrene degradation in the communities are not well-characterized.     

Substrates for Sulfate Reduction and Methane Production in Intertidal Sediments

The activity of and potential substrates for methane-producing bacteria and sulfate-reducing bacteria were examined in marsh, estuary, and beach intertidal sediments. Slow rates of methane production were detected in all sediments, although rates of sulfate reduction were 100- to 1,000-fold higher. After sulfate was depleted in sediments, the rates of methane production sharply increased. The addition of methylamine stimulated methanogenesis in the presence of sulfate, and [¹⁴C]methylamine was rapidly converted to ¹⁴CH₄ and ¹⁴CO₂ in freshly collected marsh sediment. Acetate, hydrogen, or methionine additions did not stimulate methanogenesis. [methyl-¹⁴C]methionine and [2-¹⁴C]acetate were converted to ¹⁴CO₂ and not to ¹⁴CH₄ in fresh sediment. No reduction of ¹⁴CO₂ to ¹⁴CH₄ occurred in fresh sediment. Molybdate, an inhibitor of sulfate reduction, inhibited [2-¹⁴C]acetate metabolism by 98.5%. Fluoracetate, an inhibitor of acetate metabolism, inhibited sulfate reduction by 61%. These results suggest that acetate is a major electron donor for sulfate reduction in marine sediments. In the presence of high concentrations of sulfate, methane may be derived from novel substrates such as methylamine.     

The Diverse Bacterial Community in Intertidal, Anaerobic Sediments at Sapelo Island, Georgia

The phylogenetic diversity and composition of the bacterial community in anaerobic sediments from Sapelo Island, GA, USA were examined using 16S rRNA gene libraries. The diversity of this community was comparable to that of soil, and 1,186 clones formed 817 OTUs at 99% sequence similarity. Chao1 estimators for the total richness were also high, at 3,290 OTUs at 99% sequence similarity. The program RDPquery was developed to assign clones to taxonomic groups based upon comparisons to the RDP database. While most clones could be assigned to describe phyla, fewer than 30% of the clones could be assigned to a described order. Similarly, nearly 25% of the clones were only distantly related (<90% sequence similarity) to other environmental clones, illustrating the unique composition of this community. One quarter of the clones were related to one or more undescribed orders within the γ-Proteobacteria. Other abundant groups included the δ-Proteobacteria, Bacteroidetes, and Cyanobacteria. While these phyla were abundant in other estuarine sediments, the specific members at Sapelo Island appeared to be different from those previously described in other locations, suggesting that great diversity exists between as well as within estuarine intertidal sediments. In spite of the large differences in pore water chemistry with season and depth, differences in the bacterial community were modest over the temporal and spatial scales examined and generally restricted to only certain taxa. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Predicting Epipelic Diatom Exopolymer Concentrations in Intertidal Sediments from Sediment Chlorophyll a

Abstract In many intertidal cohesive—sediment habitats, epipelic diatoms are the dominant microphytobenthic organisms. In such sediments, concentrations of colloidal carbohydrate [including the exopolymeric substances (EPS) produced by diatoms during motility] are closely correlated with the biomass (chlorophyll a) of epipelic diatoms. A model describing this relationship (log (conc. coll. carbo. + 1) = 1.40 + 1.02(log (chl. a conc. + 1)) was derived from published data. It was validated against published and unpublished data from 6 different estuaries, and accounted for 64.6% of the variation in sediment colloidal carbohydrate concentrations. The model was valid for intertidal habitats with cohesive sediments where epipelic diatoms constituted >50% of the microphytobenthic assemblage. In sites with noncohesive sediments, or where the microphytobenthic assemblage was dominated by other algal groups, the model was not applicable. The mean percentage of EPS in colloidal carbohydrate extracts varied between 11 and 37% for axenic cultures of epipelic diatoms (with higher values obtained during stationary phase), and between 22.7% and 24.3% for natural sediments dominated by epipelic diatoms. Assuming an EPS percentage of 25% in colloidal extracts yielded an EPS chl. a ratio of 2.62:1. Maximum rates of EPS production in diatom cultures occurred at the beginning of stationary phase (1.6–5.09 μg EPS μg⁻¹ chl a d⁻¹), with Nitzschia sigma having a significantly (P < 0.05) higher rate of production than N. frustulum, Navicula perminuta and Surirella ovata. Similar rates of EPS production were measured in the field. The dynamics of EPS production and loss on mudflats is discussed, with reference to the model and these production rates. Received: 25 February 1997; Accepted: 23 May 1997     

Chemoautotrophic Carbon Fixation Rates and Active Bacterial Communities in Intertidal Marine Sediments

Chemoautotrophy has been little studied in typical coastal marine sediments, but may be an important component of carbon recycling as intense anaerobic mineralization processes in these sediments lead to accumulation of high amounts of reduced compounds, such as sulfides and ammonium. We studied chemoautotrophy by measuring dark-fixation of ¹³C-bicarbonate into phospholipid derived fatty acid (PLFA) biomarkers at two coastal sediment sites with contrasting sulfur chemistry in the Eastern Scheldt estuary, the Netherlands. At one site where free sulfide accumulated in the pore water right to the top of the sediment, PLFA labeling was restricted to compounds typically found in sulfur and ammonium oxidizing bacteria. At the other site, with no detectable free sulfide in the pore water, a very different PLFA labeling pattern was found with high amounts of label in branched i- and a-PLFA besides the typical compounds for sulfur and ammonium oxidizing bacteria. This suggests that other types of chemoautotrophic bacteria were also active, most likely Deltaproteobacteria related to sulfate reducers. Maximum rates of chemoautotrophy were detected in first 1 to 2 centimeters of both sediments and chemosynthetic biomass production was high ranging from 3 to 36 mmol C m⁻² d⁻¹. Average dark carbon fixation to sediment oxygen uptake ratios were 0.22±0.07 mol C (mol O₂)⁻¹, which is in the range of the maximum growth yields reported for sulfur oxidizing bacteria indicating highly efficient growth. Chemoautotrophic biomass production was similar to carbon mineralization rates in the top of the free sulfide site, suggesting that chemoautotrophic bacteria could play a crucial role in the microbial food web and labeling in eukaryotic poly-unsaturated PLFA was indeed detectable. Our study shows that dark carbon fixation by chemoautotrophic bacteria is a major process in the carbon cycle of coastal sediments, and should therefore receive more attention in future studies on sediment biogeochemistry and microbial ecology.     

Fate of Nitrate Acquired by the Tubeworm Riftia pachyptila

The hydrothermal vent tubeworm Riftia pachyptila lacks a mouth and gut and lives in association with intracellular, sulfide-oxidizing chemoautotrophic bacteria. Growth of this tubeworm requires an exogenous source of nitrogen for biosynthesis, and, as determined in previous studies, environmental ammonia and free amino acids appear to be unlikely sources of nitrogen. Nitrate, however, is present in situ (K. Johnson, J. Childress, R. Hessler, C. Sakamoto-Arnold, and C. Beehler, Deep-Sea Res. 35:1723–1744, 1988), is taken up by the host, and can be chemically reduced by the symbionts (U. Hentschel and H. Felbeck, Nature 366:338–340, 1993). Here we report that at an in situ concentration of 40 μM, nitrate is acquired by R. pachyptila at a rate of 3.54 μmol g⁻¹ h⁻¹, while elimination of nitrite and elimination of ammonia occur at much lower rates (0.017 and 0.21 μmol g⁻¹ h⁻¹, respectively). We also observed reduction of nitrite (and accordingly nitrate) to ammonia in the trophosome tissue. When R. pachyptila tubeworms are exposed to constant in situ conditions for 60 h, there is a difference between the amount of nitrogen acquired via nitrate uptake and the amount of nitrogen lost via nitrite and ammonia elimination, which indicates that there is a nitrogen “sink.” Our results demonstrate that storage of nitrate does not account for the observed stoichiometric differences in the amounts of nitrogen. Nitrate uptake was not correlated with sulfide or inorganic carbon flux, suggesting that nitrate is probably not an important oxidant in metabolism of the symbionts. Accordingly, we describe a nitrogen flux model for this association, in which the product of symbiont nitrate reduction, ammonia, is the primary source of nitrogen for the host and the symbionts and fulfills the association's nitrogen needs via incorporation of ammonia into amino acids.     

Biogenic Stabilization of Intertidal Sediments: The Importance of Extracellular Polymeric Substances Produced by Benthic Diatoms

The sediment-stabilizing effect of benthic diatoms was investigated in a laboratory setting. Axenic cultures of the benthic diatoms Nitzschia cf. brevissima and Cylindrotheca closterium were inoculated in Petri dishes containing sand and incubated under axenic conditions. By ensuring aseptic routines throughout the experiments, interference from other organisms occurring with diatoms in natural photothrophic biofilms was avoided. This allowed the examination of the role of benthic diatoms in sediment stabilization. Increases in the critical erosion shear stress of the sediment were observed in the presence of both diatom taxa relative to sterile sediment. However, N. cf. brevissima was more effective than C. closterium. Values of critical shear stress in the experimental system were in the same range as those observed in natural biofilms, which indicates that diatoms are important agents for biogenic stabilization. Extracellular carbohydrate contents in the microcosms were similar for both diatom species. However, in the presence of N cf. brevissima, extracellular carbohydrate correlated significantly to critical shear stress, explaining up to 80% of the variation, whereas this was not the case for C. closterium. Therefore, it was concluded that the quantity of extracellular polymeric substances (EPS) alone did not explain the biogenic stabilization. Observed adsorption of EPS to sediment particles depended on the relative amount of uronic acids in the exopolymers. Using fluorescently labeled lectins, confocal laser scanning microscopy showed that EPS secretion by N. cf. brevissima resulted in ordered three-dimensional matrix structures. It is suggested that the structuring of EPS plays an prominent role in the process of biostabilization, and that diatoms such as N. cf. brevissima are actively involved in producing the structure of EPS, whereas others such as C. closterium do not do so to the same extent.     

海水养殖场沉积物中硝酸盐还原菌种群分析

通过对福建省沿海海水养殖场沉积物中参与氮循环的各生理群细菌数量分析 ,发现氨化和硝酸盐还原细菌是优势生理菌群 ,同时 ,表层泥样中的硝酸盐还原菌数量明显高于深层泥样。从该环境中分离获得 106株细菌 ,其中 58株具有硝酸盐还原能力 ,初步鉴定表明它们主要为芽孢杆菌属 (Bacillus)、盐芽孢杆菌属 (Halobacillus)、短芽孢杆菌属 (Brevibacil lus)、动性球菌属 (Planococcus)和动性杆菌属 (Plano     

Biotic Disturbance,Recolonization, and Early Succession of Bacterial Assemblages in Intertidal Sediments

The role of disturbance in structuring natural microbial communities has been largely unexplored. Disturbance associated with invertebrate ingestion can reduce bacterial biomass and alter metabolic activities and compositions of bacterial assemblages in marine sediments. The primary objectives of the research presented here were to test whether ingestion by a taxonomically diverse group of deposit feeders constituted a disturbance, and to determine the mechanisms by which bacterial assemblages recover following deposit-feeder ingestion. To test the question of disturbance, we compared fresh egesta vs surficial sediments with respect to bacterial assemblage structure. In emersed intertidal sediments, microbial recovery could be due to regrowth of bacterial populations surviving gut passage or to immigration from adjacent sediments. To differentiate between these modes of recolonization we used field manipulative experiments to exclude migration by isolating freshly extruded fecal coils of three deposit-feeding species from surrounding sediments. We then followed the quantitative and qualitative recovery in egesta and sediments through time using epifluorescence microscopy and PCR-DGGE analysis of 16S rDNA. Our findings indicate that (1) the degree and nature of the disturbance to bacterial assemblages from deposit feeding varies among invertebrate taxa, (2) recovery was significant but incomplete over 3 h, and (3) recolonization of biotically disturbed sediments is dominated by immigration.This revised version was published online in November 2004 with corrections to Volume 48.     

Stoichiometric Determination of Nitrate Fate in Agricultural Ecosystems during Rainfall Events

Ecologists have found a close relationship between the concentrations of nitrate (NO₃ ^-) and dissolved organic carbon (DOC) in ecosystems. However, it is difficult to determine the NO₃ ^- fate exactly because of the low coefficient in the constructed relationship. In the present paper, a negative power-function equation (r ² = 0.87) was developed by using 411 NO₃ ^- data points and DOC:NO₃ ^- ratios from several agricultural ecosystems during different rainfall events. Our analysis of the stoichiometric method reveals several observations. First, the NO₃ ^- concentration demonstrated the largest changes when the DOC:NO₃ ^- ratio increased from 1 to 10. Second, the biodegradability of DOC was an important factor in controlling the NO₃ ^- concentration of agricultural ecosystems. Third, sediment was important not only as a denitrification site, but also as a major source of DOC for the overlying water. Fourth, a high DOC concentration was able to maintain a low NO₃ ^- concentration in the groundwater. In conclusion, this new stoichiometric method can be used for the accurate estimation and analysis of NO₃ ^- concentrations in ecosystems.     

Microphytobenthos,their Extracellular Polymeric Substances,and the Morphogenesis of Intertidal Sediments

Intertidal sediments are important areas that separate the land from the sea and form natural coastal defenses. They are known as highly productive ecosystems, fueling the coastal food web. It is also conceived that microphytobenthos contribute to the stability of intertidal sediments by increasing the erosion threshold and that they are major players in coastal morphodynamics. Depending on the sedimentary composition of intertidal flats, different types of microphytobenthos colonize the sediment surface. Fine sand sediment is often colonized by cyanobacteria, prokaryotic algae, which form dense and rigid microbial mats. Mudflats on the other hand are characterized by the development of thin biofilms of epipelic diatoms. Both groups of phototrophic microorganisms excrete extracellular polymeric substances (EPS), but they do so in different ways and for different reasons. Two operationally defined fractions, water- and EDTA-extractable EPS, have been obtained from intertidal diatom biofilms and from cultures. They differ in composition and their production seems to be under different metabolic control. Water-extractable EPS are considered to be closely associated with the diatoms and are rich in neutral sugars, notably glucose. These EPS show a dynamic relationship with the microphytobenthic biomass. EDTA-extractable EPS are tightly bound to the sediment, probably through bridging by divalent ions. This material is rich in uronic acids and other acid sugars and is weakly related to chlorophyll. These EPS have been conceived to be a major factor in the structuring and diagenesis of coastal sediments and essential for increasing the sediment erosion threshold. However, this relationship is now questioned.     

Degradation and Mineralization of the Polycyclic Aromatic Hydrocarbons Anthracene and Naphthalene in Intertidal Marine Sediments

The degradation of the polynuclear aromatic hydrocarbons (PAHs) anthracene and naphthalene by the microbiota of intertidal sediments was investigated in laboratory studies. No mineralization of either PAH was observed in the absence of oxygen. Both rates and total amounts of PAH mineralization were strongly controlled by oxygen content and temperature of the incubations. Inorganic nitrogen and glucose amendments had minimal effects on PAH mineralization. The rates and total amounts of PAH mineralized were directly related to compound concentration, pre-exposure time, and concentration. Maximum mineralization was observed at the higher concentrations (5 to 100 μg/g [ppm]) of both PAHs. Optimal acclimation to anthracene and naphthalene (through pre-exposures to the compounds) occurred at the highest acclimation concentration (1,000 ppm). However, acclimation to a single concentration (100 ppm) resulted in initial relative mineralization rates over a range of re-exposure concentrations (1 to 1,000 ppm) being nearly identical. Maximum mineralization of both PAHs occurred after intermediate periods (1 to 2 weeks) of pre-exposure. The fraction of the total heterotrophic population capable of utilizing anthracene or naphthalene as sole carbon source was also greatest after 2 weeks.     

Influences of Infaunal Burrows on the Community Structure and Activity of Ammonia-Oxidizing Bacteria in Intertidal Sediments

Influences of infaunal burrows constructed by the polychaete (Tylorrhynchus heterochaetus) on O₂ concentrations and community structures and abundances of ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB) in intertidal sediments were analyzed by the combined use of a 16S rRNA gene-based molecular approach and microelectrodes. The microelectrode measurements performed in an experimental system developed in an aquarium showed direct evidence of O₂ transport down to a depth of 350 mm of the sediment through a burrow. The 16S rRNA gene-cloning analysis revealed that the betaproteobacterial AOB communities in the sediment surface and the burrow walls were dominated by Nitrosomonas sp. strain Nm143-like sequences, and most of the clones in Nitrospira-like NOB clone libraries of the sediment surface and the burrow walls were related to the Nitrospira marina lineage. Furthermore, we investigated vertical distributions of AOB and NOB in the infaunal burrow walls and the bulk sediments by real-time quantitative PCR (Q-PCR) assay. The AOB and Nitrospira-like NOB-specific 16S rRNA gene copy numbers in the burrow walls were comparable with those in the sediment surfaces. These numbers in the burrow wall at a depth of 50 to 55 mm from the surface were, however, higher than those in the bulk sediment at the same depth. The microelectrode measurements showed higher NH₄⁺ consumption activity at the burrow wall than those at the surrounding sediment. This result was consistent with the results of microcosm experiments showing that the consumption rates of NH₄⁺ and total inorganic nitrogen increased with increasing infaunal density in the sediment. These results clearly demonstrated that the infaunal burrows stimulated O₂ transport into the sediment in which otherwise reducing conditions prevailed, resulting in development of high NH₄⁺ consumption capacity. Consequently, the infaunal burrow became an important site for NH₄⁺ consumption in the intertidal sediment.     

Interactions between Benthic Copepods,Bacteria and Diatoms Promote Nitrogen Retention in Intertidal Marine Sediments

The present study aims at evaluating the impact of diatoms and copepods on microbial processes mediating nitrate removal in fine-grained intertidal sediments. More specifically, we studied the interactions between copepods, diatoms and bacteria in relation to their effects on nitrate reduction and denitrification. Microcosms containing defaunated marine sediments were subjected to different treatments: an excess of nitrate, copepods, diatoms (Navicula sp.), a combination of copepods and diatoms, and spent medium from copepods. The microcosms were incubated for seven and a half days, after which nutrient concentrations and denitrification potential were measured. Ammonium concentrations were highest in the treatments with copepods or their spent medium, whilst denitrification potential was lowest in these treatments, suggesting that copepods enhance dissimilatory nitrate reduction to ammonium over denitrification. We hypothesize that this is an indirect effect, by providing extra carbon for the bacterial community through the copepods'' excretion products, thus changing the C/N ratio in favour of dissimilatory nitrate reduction. Diatoms alone had no effect on the nitrogen fluxes, but they did enhance the effect of copepods, possibly by influencing the quantity and quality of the copepods'' excretion products. Our results show that small-scale biological interactions between bacteria, copepods and diatoms can have an important impact on denitrification and hence sediment nitrogen fluxes.     

Dissimilatory Nitrate Reduction in Anaerobic Sediments Leading to River Nitrite Accumulation

Recent studies on Northern Ireland rivers have shown that summer nitrite (NO(inf2)(sup-)) concentrations greatly exceed the European Union guideline of 3 (mu)g of N liter(sup-1) for rivers supporting salmonid fisheries. In fast-flowing aerobic small streams, NO(inf2)(sup-) is thought to originate from nitrification, due to the retardation of Nitrobacter strains by the presence of free ammonia. Multiple regression analyses of NO(inf2)(sup-) concentrations against water quality variables of the six major rivers of the Lough Neagh catchment in Northern Ireland, however, suggested that the high NO(inf2)(sup-) concentrations found in the summer under warm, slow-flow conditions may result from the reduction of NO(inf3)(sup-). This hypothesis was supported by field observations of weekly changes in N species. Here, reduction of NO(inf3)(sup-) was observed to occur simultaneously with elevation of NO(inf2)(sup-) levels and subsequently NH(inf4)(sup+) levels, indicating that dissimilatory NO(inf3)(sup-) reduction to NH(inf4)(sup+) (DNRA) performed by fermentative bacteria (e.g., Aeromonas and Vibrio spp.) is responsible for NO(inf2)(sup-) accumulation in these large rivers. Mechanistic studies in which (sup15)N-labelled NO(inf3)(sup-) in sediment extracts was used provided further support for this hypothesis. Maximal concentrations of NO(inf2)(sup-) accumulation (up to 1.4 mg of N liter(sup-1)) were found in sediments deeper than 6 cm associated with a high concentration of metabolizable carbon and anaerobic conditions. The (sup15)N enrichment of the NO(inf2)(sup-) was comparable to that of the NO(inf3)(sup-) pool, indicating that the NO(inf2)(sup-) was predominantly NO(inf3)(sup-) derived. There is evidence which suggests that the high NO(inf2)(sup-) concentrations observed arose from the inhibition of the DNRA NO(inf2)(sup-) reductase system by NO(inf3)(sup-).