Summary 1. Riparian zones hold a central place in the hydrological cycle, owing to the prevalence of surface and groundwater interactions. In riparian transition zones, the quality of exfiltrating water is heavily influenced by microbial activities within the bed sediments. This paper reviews the role of micro‐organisms in biogeochemical cycling in the riparian‐hyporheic ecotone. 2. The production of organic substances, such as cellulose and lignin, by riparian vegetation is an important factor influencing the pathways of microbial processing in the riparian zone. For example, anaerobic sediment patches, created by entrainment of allochthonous organic matter, are focal sites of microbial denitrification. 3. The biophysical structure of the riparian zone largely influences in‐stream microbial transformations through the retention of organic matter. Particulate and dissolved organic matter (POM and DOM) is retained effectively in the hyporheic zone, which drives biofilm development and associated microbial activity. 4. The structure of the riparian zone, the mechanisms of POM retention, the hydrological linkages to the stream and the intensity of key biogeochemical processes vary greatly along the river continuum and in relation to the geomorphic setting. However, the present state of knowledge of organic matter metabolism in the hyporheic zone suggests that lateral ecological connectivity is a basic attribute of lotic ecosystems. 5. Due to their efficiency in transforming POM into heterotrophic microbial biomass, attached biofilms form an abundant food resource for an array of predators and grazers in the interstitial environments of rivers and streams. The interstitial microbial loop, and the intensity of microbial production within the bed sediments, may be a primary driver of the celebrated high productivity and biodiversity of the riparian zone. 6. New molecular methods based on the analysis of the low molecular weight RNA (LMW RNA) allow unprecedented insights into the community structure of natural bacterial assemblages and also allow identification and study of specific strains hitherto largely unknown. 7. Research is needed on the development and evaluation of sampling methods for interstitial micro‐organisms, on the characterization of biofilm structure, on the analysis of the biodegradable matter in the riparian‐hyporheic ecotone, on the regulation mechanisms exerted on microbiota by interstitial predators and grazers, and on measures of microbial respiration and other key activities that influence biogeochemical cycles in running waters. 8. Past experiences from large‐scale alterations of riparian zones by humans, such as the River Rhine in central Europe, undeniably demonstrate the detrimental consequences of disconnecting rivers from their riparian zones. A river management approach that uses the natural services of micro‐organisms within intact riparian zones could substantially reduce the costs of clean, sustainable water supplies for humans.

Biostimulation is widely used to enhance reductive dechlorination of chlorinated ethenes in contaminated aquifers. However, the knowledge on corresponding biogeochemical responses is limited. In this study, glycerol was injected in an aquifer contaminated with cis-dichloroethene (cDCE), and geochemical and microbial shifts were followed for 265 days. Consistent with anoxic conditions and sulfate reduction after biostimulation, MiSeq 16S rRNA gene sequencing revealed temporarily increased relative abundance of Firmicutes, Bacteriodetes and sulfate reducing Deltaproteobacteria. In line with C cDCE enrichment and increased Dehalococcoides mccartyi (Dcm) numbers, dechlorination was observed toward the end of the field experiment, albeit being incomplete with accumulation of vinyl chloride. This was concurrent with (i) decreased concentrations of dissolved organic carbon (DOC), reduced relative abundances of fermenting and sulfate reducing bacteria that have been suggested to promote Dcm growth by providing electron donor (H ) and essential corrinoid cofactors, (ii) increased sulfate concentration and increased relative abundance of Epsilonproteobacteria and Deferribacteres as putative oxidizers of reduced sulfur compounds. Strong correlations of DOC, relative abundance of fermenters and sulfate reducers, and dechlorination imply the importance of syntrophic interactions to sustain robust dechlorination. Tracking microbial and environmental parameters that promote/preclude enhanced reductive dechlorination should aid development of sustainable bioremediation strategies.

A pilot injection test with guar gum stabilized microscale zerovalent iron (mZVI) particles was performed at test site V (Belgium) where different chlorinated aliphatic hydrocarbons (CAHs) were present as pollutants in the subsurface. One hundred kilograms of 56μm-diameter mZVI (~70gL(-1)) was suspended in 1.5m(3) of guar gum (~7gL(-1)) solution and injected into the test area. In order to deliver the guar gum stabilized mZVI slurry, one direct push bottom-up injection (Geoprobe) was performed with injections at 5 depths between 10.5 and 8.5m bgs. The direct push technique was preferred above others (e.g. injection at low flow rate via screened wells) because of the limited hydraulic conductivity of the aquifer, and to the large size of the mZVI particles. A final heterogeneous distribution of the mZVI in the porous medium was observed explicable by preferential flow paths created during the high pressure injection. The maximum observed delivery distance was 2.5m. A significant decrease in 1,1,1-TCA concentrations was observed in close vicinity of spots where the highest concentration of mZVI was observed. Carbon stable isotope analysis (CSIA) yielded information on the success of the abiotic degradation of 1,1,1-TCA and indicated a heterogeneous spatio-temporal pattern of degradation. Finally, the obtained results show that mZVI slurries stabilized by guar gum can be prepared at pilot scale and directly injected into low permeable aquifers, indicating a significant removal of 1,1,1-TCA.

Confocal laser scanning microscopy (CLSM) is an extensive but reliable tool for assessing the hybridisation signals in fluorescence in situ hybridisation (FISH). Most CLSMs are equipped with an argon-laser and a helium/neon-laser illumination system with excitation wavelengths of 488, 543 and 633 nm. A protocol for an optimal nuclear counterstaining in combination with dual-colour FISH for these laser illumination systems has not been established so far. Here, we determined the suitability of eleven dimeric and monomeric cyanine nucleic acid stains on paraffin sections of breast carcinoma specimens in combination with dual-colour FISH (Her-2/neu and centromere 17) for CLSM application. Strong staining of cell nuclei was observed for TO-PRO-3 and YO-PRO-3, YOYO-1 and propidium iodide (PI), but only TO-PRO-3 showed specific staining of nuclei without any staining of the cytoplasm. A specific emission in exclusively one distinct fluorescence channel was shown for TO-PRO-3 (633 nm excitation) as well as YOYO-1, BO-PRO-1 and Sytox Green (488 nm excitation), evaluated by a CLSM and confirmed by 3-D fluorescence spectra. High stability of fluorescence intensity was shown for the far-red dyes TO-PRO-3, YO-PRO-3, YOYO-3 and Syto-59 as well as YOYO-1 and PI. Only TO-PRO-3 was due to its high specificity and stability suitable for detection of an amplification of the Her-2/neu gene by dual-colour FISH and CLSM evaluation.

We investigated attachment processes of hydrophobic and hydrophilic particles (diameter ‫؍‬ 1 m) to mature biofilms grown on clay marbles in a sequencing batch biofilm reactor. During a treatment cycle with filtered wastewater containing different fluorescent beads, the progression of particle density in various biofilm compartments (carrier biofilm, basic biofilm layer, biofilm flocs, and sessile ciliates) was determined by flow cytometry, confocal laser scanning microscopy and automated image analysis. Particles were almost completely removed from wastewater by typical processes of particle retention: up to 58% of particles attached to clay marbles, up to 15% were associated with suspended flocs, and up to 10% were ingested by sessile ciliates. Ingestion of particles by ciliates was exceptionally high immediately after wastewater addition (1,200 particles grazer ؊1 h ؊1 ) and continued until approximately 14% of the water had been cleared by ciliate filter feeding. Most probably, ciliate bioturbation increases particle sorption to the basic biofilm. Backwashing of the reactor detached pieces of biofilm and thus released approximately 50% of the particles into rinsing water. Clay marbles in the upper part of the reactor were more efficiently abraded than in the lower part. No indications for selective attachment of the applied hydrophobic and hydrophilic beads were found. As a consequence of interception patterns, organisms at elevated biofilm structures are probably major profiteers of wastewater particles; among them, ciliates may be of major importance because of their highly active digestive food vacuoles.