BackgroundMicrobial denitrification is not considered important in human-associated microbial communities. Accordingly, metabolic investigations of the microbial biofilm communities of human dental plaque have focused on aerobic respiration and acid fermentation of carbohydrates, even though it is known that the oral habitat is constantly exposed to nitrate (NO3-) concentrations in the millimolar range and that dental plaque houses bacteria that can reduce this NO3- to nitrite (NO2-).ResultsWe show that dental plaque mediates denitrification of NO3- to nitric oxide (NO), nitrous oxide (N2O), and dinitrogen (N2) using microsensor measurements, 15N isotopic labelling and molecular detection of denitrification genes. In vivo N2O accumulation rates in the mouth depended on the presence of dental plaque and on salivary NO3- concentrations. NO and N2O production by denitrification occurred under aerobic conditions and was regulated by plaque pH.ConclusionsIncreases of NO concentrations were in the range of effective concentrations for NO signalling to human host cells and, thus, may locally affect blood flow, signalling between nerves and inflammatory processes in the gum. This is specifically significant for the understanding of periodontal diseases, where NO has been shown to play a key role, but where gingival cells are believed to be the only source of NO. More generally, this study establishes denitrification by human-associated microbial communities as a significant metabolic pathway which, due to concurrent NO formation, provides a basis for symbiotic interactions.

Summary Emission of the greenhouse gas nitrous oxide (N2O) from freshwater and terrestrial invertebrates has exclusively been ascribed to N2O production by ingested denitrifying bacteria in the anoxic gut of the animals. Our study of marine molluscs now shows that also microbial biofilms on shell surfaces are important sites of N2O production. The shell biofilms of Mytilus edulis, Littorina littorea and Hinia reticulata contributed 18–94% to the total animal‐associated N2O emission. Nitrification and denitrification were equally important sources of N2O in shell biofilms as revealed by 15N‐stable isotope experiments with dissected shells. Microsensor measurements confirmed that both nitrification and denitrification can occur in shell biofilms due to a heterogeneous oxygen distribution. Accordingly, ammonium, nitrite and nitrate were important drivers of N2O production in the shell biofilm of the three mollusc species. Ammonium excretion by the animals was found to be sufficient to sustain N2O production in the shell biofilm. Apparently, the animals provide a nutrient‐enriched microenvironment that stimulates growth and N2O production of the shell biofilm. This animal‐induced stimulation was demonstrated in a long‐term microcosm experiment with the snail H. reticulata, where shell biofilms exhibited the highest N2O emission rates when the animal was still living inside the shell.

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ABSTRACTNitrification in shell biofilms and denitrification in the gut of the animal accounted for N2O emission byDreissena polymorpha(Bivalvia), as shown by gas chromatography and gene expression analysis. The mussel's ammonium excretion was sufficient to sustain N2O production and thus potentially uncouples invertebrate N2O production from environmental N concentrations.

In an intertidal flat of the German Wadden Sea, a large sedimentary pool of intracellular nitrate was discovered that by far exceeded the pool of nitrate that was freely dissolved in the porewater. Intracellular nitrate was even present deep in anoxic sediment layers where it might be used for anaerobic respiration processes. The origin and some of the ecological controls of this intracellular nitrate pool were investigated in a laboratory experiment. Sediment microcosms were set up with and without the abundant polychaete Hediste diversicolor that is known to stimulate nitrate production by microbial nitrification in the sediment. Additional treatments were amended with ammonium to mimic ammonium excretion by the worms or with allylthiourea (ATU) to inhibit nitrification by sediment bacteria. H. diversicolor and ammonium increased, while ATU decreased the intracellular nitrate pool in the sediment. Microsensor profiles of porewater nitrate showed that bacterial nitrification was enhanced by worms and ammonium addition. Thus, nitrification formed an important nitrate supply for the intracellular nitrate pool in the sediment. The vertical distribution of intracellular nitrate matched that of the photopigments chlorophyll a and fucoxanthin, strongly suggesting that diatoms were the main nitrate-storing organisms. Intracellular nitrate formation is thus stimulated by the interaction of phylogenetically distant groups of organisms: worms enhance nitrification by feeding on particulate organic matter, excreting ammonium and oxygenating the sediment; bacteria oxidise ammonium to nitrate in oxic sediment layers; and diatoms store nitrate intracellularly. KEY WORDS: Intertidal sediment · Wadden Sea · Nitrogen cycle · Intracellular nitrate · Microphytobenthos · Macrofauna · Nereis diversicolor · Trophic interaction Resale or republication not permitted without written consent of the publisherMar Ecol Prog Ser 445: [181][182][183][184][185][186][187][188][189][190][191][192] 2012 and probably also intracellular nitrate for nitrogen assimilation (Lomas & Glibert 2000, Sundbäck & Miles 2000.The ability to store nitrate intracellularly may be particularly advantageous in intertidal flats, which are very dynamic ecosystems. Benthic organisms must cope with frequent changes in the availability of e.g. light, oxygen and nutrients due to tidal and diurnal rhythms. Another source of perturbation in intertidal flats is the presence of macrofauna that rework large amounts of sediment and the microorganisms therein (e.g. Bouchet et al. 2009). Some polychaetes construct deep-reaching burrows and enhance solute exchange between sediment and the water column due to their ventilation activity (Kristensen 2001). Many species of intertidal macrofauna feed on sediment microorganisms and thereby decrease microbial populations or keep them in the exponential growth phase (Herman et al. 2000, Blanchard et al. 2001). Under such transient conditions, the nitrate storage capacity awards sediment microorganisms with the steady availa...