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NO 3 − concentration profiles were measured in the sediments of a meso-eutrophic lake with a newly developed microelectrode. The depth of penetration of NO 3 − varied from only 1.3 mm in organic-rich profundal silty sediments to 5 mm in organic-poor littoral sandy sediments. The thickness of the zone of denitrification in the organic-rich sediments was −2 day −1 ) at in situ nitrate concentrations in the overlying water (0.030 mmol liter −1 ) and limitation of denitrification by nitrate availability.

In this article, we describe the dynamics of pH, O2 and H2S in the top 5–10 cm of an intertidal flat consisting of permeable sand. These dynamics were measured at the low water line and higher up the flat and during several seasons. Together with pore water nutrient data, the dynamics confirm that two types of transport act as driving forces for the cycling of elements (Billerbeck et al. 2006b): Fast surface dynamics of pore water chemistry occur only during inundation. Thus, they must be driven by hydraulics (tidal and wave action) and are highly dependent on weather conditions. This was demonstrated clearly by quick variation in oxygen penetration depth: Seeps are active at low tide only, indicating that the pore water flow in them is driven by a pressure head developing at low tide. The seeps are fed by slow transport of pore water over long distances in the deeper sediment. In the seeps, high concentrations of degradation products such as nutrients and sulphide were found, showing them to be the outlets of deep-seated degradation processes. The degradation products appear toxic for bioturbating/bioirrigating organisms, as a consequence of which, these were absent in the wider seep areas. These two mechanisms driving advection determine oxygen dynamics in these flats, whereas bioirrigation plays a minor role. The deep circulation causes a characteristic distribution of strongly reduced pore water near the low water line and rather more oxidised sediments in the centre of the flats. The two combined transport phenomena determine the fluxes of solutes and gases from the sediment to the surface water and in this way create specific niches for various types of microorganisms.

The Mediterranean sponge Aplysina aerophoba kept in aquaria or cultivation tanks can stop pumping for several hours or even days. To investigate changes in the chemical microenvironments, we measured oxygen profiles over the surface and into the tissue of pumping and non-pumping A. aerophoba specimens with Clark-type oxygen microelectrodes (tip diameters 18-30 μm). Total oxygen consumption rates of whole sponges were measured in closed chambers. These rates were used to back-calculate the oxygen distribution in a finite-element model. Combining direct measurements with calculations of diffusive flux and modeling revealed that the tissue of non-pumping sponges turns anoxic within 15 min, with the exception of a 1 mm surface layer where oxygen intrudes due to molecular diffusion over the sponge surface. Molecular diffusion is the only transport mechanism for oxygen into non-pumping sponges, which allows total oxygen consumption rates of 6-12 μmol cm-3 sponge day-1. Sponges of different sizes had similar diffusional uptake rates, which is explained by their similar surface/volume ratios. In pumping sponges, oxygen consumption rates were between 22 and 37 μmol cm-3 sponge day-1, and the entire tissue was oxygenated. Combining different approaches of direct oxygen measurement in living sponges with a dynamic model, we can show that tissue anoxia is a direct function of the pumping behavior. The sponge-microbe system of A. aerophoba thus has the possibility to switch actively between aerobic and anaerobic metabolism by stopping the water flow for more than 15 min. These periods of anoxia will greatly influence physiological variety and activity of the sponge microbes. Detailed knowledge about the varying chemical microenvironments in sponges will help to develop protocols to cultivate sponge-associated microbial lineages and improve our understanding of the sponge-microbe-system.

Nitrate-selective microelectrodes (1-μm tip diameter) with a liquid membrane ion exchanger were prepared. The response of the electrode is linear from 10 −1 to 10 −5 M nitrate and the response time is 30 s. Selectivity coefficients are of the same order as specified for macroelectrodes. Measurements in sediments of a mesotrophic lake and measurements in spherical agar gels with immobilized denitrifying bacteria are presented.

Ammonium-selective microelectrodes made with an ion-exchanger based on nonactin and having a tip diameter of 1 mum have been developed. The response of the electrode is linear from 10(-5) to 10(-1)M NH(+)(4) and the response time is 1 min. Applications of these electrodes can be found in biotechnology and microbial ecology, but the low selectivity with respect to K(+) and Na(+) limits their use to low salt environments. Concentration gradients in gels containing cross-linked urease were measured and found to be in accord with macrokinetic measurements.

Noninvasive H-1-nuclear magnetic resonance (NMR) imaging was used to investigate the diffusive properties of microbial mats in two dimensions. Pulsed field gradient NMR was used to acquire images of the H2O diffusion coefficient, D-s,D- and multiecho imaging NMR was used to obtain images of the water density in two structurally different microbial mats sampled from Solar Lake (Egypt). We found a pronounced lateral and vertical variability of both water density and water diffusion coefficient, correlated with the laminated and heterogeneous distribution of microbial cells and exopolymers within the mats. The average water density varied from 0.5 to 0.9, whereas the average water diffusion coefficient ranged from 0.4 to 0.9 relative to the values obtained in the stagnant water above the mat samples. The apparent water diffusivities estimated from NMR imaging compared well to apparent O-2 diffusivities measured with a diffusivity microsensor. Analysis of measured O-2 concentration profiles with a diffusion-reaction model showed that both the magnitude of calculated rates and the depth distribution of calculated O-2 consumption/production zones changed when the observed variations of diffusivity were taken into account. With NMR imaging, diffusivity can be determined at high spatial resolution, which can resolve inherent lateral and vertical heterogeneities found in most natural benthic systems.

An ammonium ion-selective microelectrode is described. With this microelectrode, product gradients were measured in an enzymic model system consisting of cross-linked urease immobilized in spherical agar gels. The measured and calculated gradients showed a good correspondence. pH gradients measured in a microbial system of aggregated methanogens diverged significantly from those calculated with a structured mathematical model. It is concluded that microelectrodes are reliable and are of great value for the understanding of heterogeneous biological systems.

granules suspended ina mineral mediumlacking glucose exhibited a distinct internal pHdecrease ofabout1U within thegranule, suggesting strong metabolism bytheacidogenic bacteria. Molasses-acclimatized andagedgranules suspended inmineral mediumdidnotexhibit such apHdecrease, suggesting theimportance ofthemetabolic state ofthese acidogens. ThepHgradient didnotoccurindeactivated granules andwas notobservable in strongly buffered media(mineral mediumcontaining 33mM phosphate or reactor liquid). Whenglucose (0.5 to5.0mM) was addedtothemineral medium, granules exhibited a convexpHprofile. Glucose consumption was located exclusively intheouter200to300 .moftheaggregates (meandiameter = 1.5mm).Theaddition of20mM 2-bromoethanesulfonic acidtothemineral mediumindicated that thehigher pHlevels inthecentre ofthegranule appeared toberelated totheactivity ofmethanogens. Itissuggested thatacidogenic activity occurspredominantly intheouter200to300,umoftheaggregate andmethanogenic activity occurs predominantly inthecenter oftheinvestigated granules. Themicrobial community involved inanaerobic digestion processes innatural habitats aswellasinmanufactured digestion systems isknowntobequite complex, comprising hydrolytic, fermentative, acidogenic, andmethanogenic bacteria. Complete anaerobic degradation oforganic matter to methane (CH4)andcarbon dioxide (CO2) requires theconcerted action ofthese different microbial species (4,13,63). Anaerobic wastewater treatment relies onthecontrol of these microbial interactions byapplying appropriate reactor conditions andreactor configurations (22, 40). Theupflow anaerobic sludge blanket (UASB)reactor concept (29, 30) uncouples solid andhydraulic retention times bybiomass accumulation incompact, well-settling aggregates. This granular configuration hasseveral engineering advantages suchasamaximal microorganism-to-space ratio andfacili

This paper presents a new non-invasive technique for measuring sediment O2 uptake that, in its concept, differs fundamentally from other methods used to date. In almost all natural aquatic environments, the vertical transport of O2 through the water column toward the sediment surface is facilitated by turbulent motion. The new technique relies on measuring 2 parameters simultaneously and at the same point in the water above the sediment: the fluctuating vertical veloc- ity using an acoustic Doppler velocimeter and the fluctuating O2 concentration using an O2 micro- electrode. From these 2 parameters, which typically are measured 10 to 50 cm above the sediment surface for a period of 10 to 20 min and at a frequency of 15 to 25 Hz, the vertical flux of O2 toward the sediment surface is derived. Based on measurements performed under actual field conditions and comparisons with in situ flux-chamber measurements, we believe that this new technique is the opti- mal approach for determining O2 uptake by sediments. The technique is superior to conventional methods as measurements are done under true in situ conditions, i.e. without any disturbance of the sediment and under the natural hydrodynamic conditions. Furthermore, this technique can be used for bio-irrigated or highly permeable sediments, such as sands, where traditional methods often fail. While this paper only focuses on O2 uptake by sediments, the technique can also be applied to other solutes that can be measured at a sufficiently high temporal resolution.

During the night of 22-23 October 2002, a large amount of airborne dust fell with rain over Canberra, located some 200 km from Australia's east coast, and at an average altitude of 650 m. It is estimated that during that night about 6 g m-2 of aeolian dust fell. We have conducted a vast number of analyses to "fingerprint" some of the dust and used the following techniques: grain size analysis; scanning electron microscope imagery; major, trace, and rare earth elemental, plus Sr and Nd isotopic analyses; organic compound analyses with respective compound-specific isotope analyses; pollen extraction to identify the vegetation sources; and molecular cloning of 16S rRNA genes in order to identify dust bacterial composition. DNA analyses show that most obtained 16S rRNA sequences belong mainly to three groups: Proteobacteria (25%), Bacteriodetes (23%), and gram-positive bacteria (23%). In addition, we investigated the meteorological conditions that led to the dust mobilization and transport using model and satellite data. Grain sizes of the mineral dust show a bimodal distribution typical of proximal dust, rather than what is found over oceans, and the bimodal aspect of size distribution confirms wet deposition by rain droplets. The inorganic geochemistry points to a source along/near the Darling River in NW New South Wales, a region that is characteristically semiarid, and both the organic chemistry and palynoflora of the dust confirm the location of this source area. Meteorological reconstructions of the event again clearly identify the area near Bourke-Cobar as being the source of the dust. This study paves the way for determining the export of Australian airborne dust both in the oceans and other continents.

The effect of sediment-dwelling macrofauna on nitrifying bacteria was investigated by direct probing in their overlapping habitat, i.e., the upper few millimeters of freshwater sediments. Microsensors (O2 ,N H þ ,N O 2 , and diffusivity) were used at the sediment surface and inside animal burrows to record steady-state and dynamic distributions of reactants, respectively. Short-term changes of metabolic activity (actual and potential nitrification rates) and long-term changes of abundance (fluorescence in situ hybridization) of nitrifying bacteria were determined. The presence of insect larvae (Chironomus riparius) increased the availability of O2 and NO 2 in the sediment pore water and inside animal burrows, suggesting promotion of nitrification and dissimilatory NO 2 reduction, particularly in the burrowing layer of C. riparius. At the sediment surface (i.e., in the feeding layer of C. riparius), however, nitrification was inhibited by low NH þ availability and high macrofaunal grazing pressure. Consequently, both actual and potential nitrification rates decreased in the feeding layer. Inside burrows, no net nitrification was detected, despite high NH þ availability and frequent O2 injections by larval ventilation activity. Conversely, burrows were sites of NH þ production and NO 2 consumption. Nevertheless, the abundance of nitrifying bacteria increased measurably in the burrowing layer after prolonged incubation, but only in sediments in which the larvae were able to construct and ventilate stable burrows.

Using molecular techniques and microsensors for H 2 S and CH 4 , we studied the population structure of and the activity distribution in anaerobic aggregates. The aggregates originated from three different types of reactors: a methanogenic reactor, a methanogenic-sulfidogenic reactor, and a sulfidogenic reactor. Microsensor measurements in methanogenic-sulfidogenic aggregates revealed that the activity of sulfate-reducing bacteria (2 to 3 mmol of S 2− m −3 s −1 or 2 × 10 −9 mmol s −1 per aggregate) was located in a surface layer of 50 to 100 μm thick. The sulfidogenic aggregates contained a wider sulfate-reducing zone (the first 200 to 300 μm from the aggregate surface) with a higher activity (1 to 6 mmol of S 2− m −3 s −1 or 7 × 10 −9 mol s −1 per aggregate). The methanogenic aggregates did not show significant sulfate-reducing activity. Methanogenic activity in the methanogenic-sulfidogenic aggregates (1 to 2 mmol of CH 4 m −3 s −1 or 10 −9 mmol s −1 per aggregate) and the methanogenic aggregates (2 to 4 mmol of CH 4 m −3 s −1 or 5 × 10 −9 mmol s −1 per aggregate) was located more inward, starting at ca. 100 μm from the aggregate surface. The methanogenic activity was not affected by 10 mM sulfate during a 1-day incubation. The sulfidogenic and methanogenic activities were independent of the type of electron donor (acetate, propionate, ethanol, or H 2 ), but the substrates were metabolized in different zones. The localization of the populations corresponded to the microsensor data. A distinct layered structure was found in the methanogenic-sulfidogenic aggregates, with sulfate-reducing bacteria in the outer 50 to 100 μm, methanogens in the inner part, and Eubacteria spp. (partly syntrophic bacteria) filling the gap between sulfate-reducing and methanogenic bacteria. In methanogenic aggregates, few sulfate-reducing bacteria were detected, while methanogens were found in the core. In the sulfidogenic aggregates, sulfate-reducing bacteria were present in the outer 300 μm, and methanogens were distributed over the inner part in clusters with syntrophic bacteria.

The most recent checklist of mosses in Belgium, dating from 2007, indicated for the first time the region in which they occur. As far as Flanders is concerned, this list has been critically reviewed. 32 species were added because they were either discovered after the publication of the checklist, or because they were inadvertently not included in the checklist. 27 species were deleted because of lack of evidence or because they were wrongly identified. The adapted checklist in annex has been supplemented with a rarity indicator and the Red List status of these species in Flanders.

Gradients of oxygen and sulfide measured towards individual cells of the large nitrate-storing sulfur bacterium Thiomargarita namibiensis showed that in addition to nitrate oxygen is used for oxidation of sulfide. Stable gradients around the cells were found only if acetate was added to the medium at low concentrations.

Summary Local nitrification and carbon assimilation activities were studied in situ in a model biofilm to investigate carbon yields and contribution of distinct populations to these activities. Immobilized microcolonies (related to Nitrosomonas europaea / eutropha , Nitrosomonas oligotropha , Nitrospira sp., and to other Bacteria ) were incubated with [ 14 C]-bicarbonate under different experimental conditions. Nitrifying activity was measured concomitantly with microsensors (oxygen, ammonium, nitrite, nitrate). Biofilm thin sections were subjected to fluorescence in situ hybridization (FISH), microautoradiography (MAR), and local quantification of [ 14 C]-bicarbonate uptake (beta microimaging). Nitrifying activity and tracer assimilation were restricted to a surface layer of different thickness in the various experiments (substrate or oxygen limitation). Excess oxygen uptake under all conditions revealed heterotrophic activity fuelled by decay or excretion products during active nitrification. Depth limits and intensity of tracer incorporation profiles were in agreement with ammonia-oxidation activity (measured with microsensors), and distribution of incorporated tracer (detected with MAR). Microautoradiography revealed a sharp individual response of distinct populations in terms of in-/activity depending on the (local) environmental conditions within the biofilm. Net in situ carbon yields on N, expressed as e ‐ equivalent ratios, varied between 0.005 and 0.018, and, thus, were in the lower range of data reported for pure cultures of nitrifiers.

We investigated the hypothesis that sulfate reduction rather than oxygenic photosynthesis promotes calcification in a hypersaline microbial mat by increasing the ion concentration product: ICP 5 [Ca 21 ] 3 [CO ]. Pore-water 22 3 calcium concentration profiles directly measured with microsensors show that calcium concentration in the photic zone decreased in illuminated mats and increased slightly in dark mats. High pH values in the photic zone of illuminated mats resulted in higher carbonate concentrations (2.25 mmol L 21 ) than in dark mats (0.75 mmol L 21 ), although the dissolved inorganic carbon (DIC) pore-water concentration in the former was much lower (5.9 mmol L 21 ) than in the latter (9.9 mmol L 21 ). The pH-induced rise in carbonate concentration in the light was the main factor influencing the ICP, while changes in Ca 21 concentration played a subsidiary role. Sulfate reduction did not result in a net pH increase in these mats, as rates in the photic zone were comparable between illuminated and dark mats (4 and 5 nmol cm 22 h 21 , respectively), and pH increased in illuminated mats but not in dark mats. Calcium carbonate precipitation in the photic zone of these hypersaline mats is primarily controlled by photosynthesisinduced pH and carbonate concentration increases. However, heterotrophic bacteria, including sulfate reducers, play an important complementary role in calcification because they maintain high concentrations of DIC in the mat pore water.

Nitrification was studied with microsensors and fluorescence in situ hybridization (FISH) in sandy sediment of a small lowland stream. Comparative measurements were performed in both intact field sediment ('natural sediment') and sediment from the same site after processing for a laboratory incubation experiment ('manipulated sediment'). In natural sediment, the nitrification activity and abundance of nitrifiers were markedly low. In contrast, nitrification activity in manipu- lated sediment (sieved, homogenized, and incubated for 5 wk in the laboratory with NH4 + -enriched stream water) was significantly higher. Similarly, abundances of NH4 + -oxidizing β-proteobacteria (AOB) and NO2 - -oxidizing Nitrospira spp. directly at the sediment surface were markedly higher than in natural sediment. AOB mixed into deep sediment layers by homogenization disappeared more quickly than Nitrospira spp., suggesting that the latter were more persistent under anoxic con- ditions. Higher activity and abundance of nitrifiers near the sediment -water interface of manipulated sediment were explained by (1) the additional NH4 + supply via the overlying water and (2) the adverse conditions for nitrification in the field. In conclusion, the snapshot measurement in natural sediment revealed the spatial heterogeneity created by stream dynamics, whereas the sediment manipulation provided semi-natural microcosms with reduced heterogeneity suitable for factorial experiments.

The coupling between CO2 and Ca2+ exchange and photosynthesis by corals (Favia sp.) was studied with microsensors for Ca2+, O2, pH and CO2. The profiles of these compounds, measured perpendicular on the coral surface, were strongly influenced by light. During illumination, the concentration of O2 and the pH at the polyp surface was higher than in the surrounding seawater, while the concentrations of Ca2+ and CO2 were lower. In the dark the inverse was observed. Furthermore, simultaneous recording of concentration changes at the coral surface, in response to light and inhibitors, were performed with pairs of the sensors. The concentration changes of CO2 and pH were slow, while those of Ca2+ and O2 were immediate and fast. The concentration changes of the O2 and Ca2+ concentrations at the coral surface were synchronous in response to changes in light conditions and to inhibition of the photosynthesis. Also, the spatial distribution of photosynthetic activity over a single polyp coincided with the distribution of Ca2+ concentration changes. These results show that Ca2+ dynamics at the polyp surface is not an indirect effect of increased CaCO3 precipitation at the skeleton, but indicates the presence of a Ca2+ uptake mechanism that is directly correlated to photosynthesis. Inhibition of carbonic anhydrase strongly decreased photosynthesis, especially at higher light intensities. This, combined with the observed increase in CO2 concentration changes and absolute increase in CO2 concentration at the tissue surface, demonstrated the importance of carbonic anhydrase for CO2/DIC uptake and transport to the site of photosynthesis.

The archaeal communities of six anaerobic granular biofilm samples were surveyed for the occurrence of Crenarchaeota, using clone library and terminal restriction fragment length polymorphism (TRFLP) analysis. One of the samples, containing a high abundance of Crenarchaeota-like organisms, was used to inoculate anaerobic reactors for the treatment of a whey-based wastewater. TRFLP was used to monitor the Crenarchaeota-like species during the trial in response to various perturbations. FISH was employed to establish the spatial arrangement of Archaea, Crenarchaeota and Bacteria within biofilm architecture. Radioactive tracer experiments and micro-beta-imaging were used to describe substrate uptake patterns by members of the biofilm. The work represents a groundbreaking effort to investigate the role and in situ functionalism of Crenarchaeota and other groups within biotechnologically important granular biofilm-based ecosystems.

From the Hakon Mosby Mud Volcano (HMMV) on the southwest Barents Sea shelf, gas and fluids are expelled by active mud volcanism. We studied the mass transfer phenomena and microbial conversions in the surface layers using in situ microsensor measurements and on retrieved cores. The HMMV consists of three concentric habitats: a central area with gray mud, a surrounding area covered by white mats of big sulfide oxidizing filamentous bacteria (Beggiatoa), and a peripheral area colonized by symbiontic tube worms (Pogonophora). A fourth habitat comprised gray microbial mats near gas seeps. The differences between these four methane-fueled habitats are best explained by different transport rates of sulfate into the sediments and pore- water upflow rates. The upflow velocities were estimated by two independent methods at 3-6 m yr21 in the central area and 0.3-1 m yr21 in Beggiatoa mats. In the central area no sulfide was found, indicating that the rapidly rising sulfate-free fluids caused sulfate limitation that inhibited anaerobic oxidation of methane (AOM). Under Beggiatoa mats a steep sulfide peak was found at 2 to 3 cm below the seafloor (bsf), most likely due to AOM. All sulfide was oxidized anaerobically, possibly through nitrate reduction by Beggiatoa. The Beggiatoa mats were dominated by a single filamentous morphotype with a diameter of 10 mm and abundant sulfur inclusions. A high diversity of sulfide oxidizer morphotypes was observed in a grayish microbial mat near gas vents, where aerobic sulfide oxidation was important. The sediments colonized by Pogonophora were influenced by bioventilation, allowing sulfate penetration and AOM to 70 cm bsf. The HMMV is a unique and diverse ecosystem, the structure and functioning of which is mainly controlled by pore-water flow.

A new CO, microelectrode with a tip diameter of 10 pm and a response time (t,,) of -10 s is presented. The sensor allows CO, measurements with a detection limit of

A glucose oxidase-based needle-type microsensor is described that is independent of the local oxygen concentration. The sensor consists of an enzyme-coated platinum microelectrode that is inserted in a glass capillary with a tip of 5–25 μm, and connected to the environment via an agar membrane. Oxygen is supplied to the enzyme coating from the shaft of the capillary. Depending on the electrode configuration, the sensor has a response time of 1–10 s, and the measuring range extends from 0.01 to 0.4 mmol l −1 or from 0.4 to 10 mmol l −1 . The signal is not affected by stirring.

We used microsensors to study short-term temperature effects on gross photosynthesis, oxygen consumption and sulfate reduction in an extremely hypersaline (salinity 200‰) cyanobac- terial mat. In situ, the mat surface can reach high temperatures (ca. 50°C in summer), and experi- ences daily temperature fluctuations due to tidal inundation. A mat sample was incubated in the lab- oratory at 25, 35, 45, 50 and 60°C. Areal rates of gross and net photosynthesis increased with increasing temperature, and maximum rates were detected at 45°C. The photosynthetic zone (i.e. the depth zone in which oxygenic photosynthesis was detectable by the light-dark shift method) decreased from 3 to 1.75 mm with increasing temperature. Above 50°C, photosynthesis was com- pletely inhibited, probably due to high sulfide concentrations. This inhibition was confirmed by the pH profiles, where the peak in the top 2 to 4 mm at 25 to 45°C disappeared at 50 to 60°C. Between 25 and 45°C, areal oxygen consumption rates in the light did not change significantly but showed a decreasing trend in the photosynthetic zone, suggesting a decoupling between photosynthesis and oxygen consumption in the light. In both the light and dark, sulfide production was strongly en- hanced by increasing temperature to a maximum flux at 50°C, above which it became inhibited. At 50 to 60°C, the sulfide produced could not be completely removed by sulfide oxidation, resulting in sulfide leakage to the overlying water. Our data demonstrate that these mat ecosystems are well adapted to high ambient temperatures. Photosynthesis, oxygen consumption in the light and sulfide production are differentially controlled by temperature. However, the individual processes in this complex community are not only directly controlled by temperature, but also by the temperature- dependent response of other processes.