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Hyporheic zones are considered hot spots for numerically vast and phylogenetically diverse microbial communities. However, biogeochemical effects of hyporheic zones have rarely been investigated in detail because of the difficulty in accurately measuring denitrification in these zones. To date, little is known about the hydroecology of hyporheic zones. The effect of changes in hydraulic conditions on the community variations of indigenous microorganisms and water quality was examined based on the depth of the hyporheic zone. In particular, we report on the use of the pyrosequencing technique to elucidate denitrifying bacteria (DNB) community profiles combined with the stable isotope composition of nitrate and hydrological patterns in the hyporheic zones to reveal whether denitrification occurs. δ 15 N–NO 3 and δ 18 O–NO 3 values of nitrate were analyzed to evaluate the transformation processes of nitrate in upwelling and downwelling areas and mixed zones. The isotope values indicated different origins of water in upwelling and downwelling zones and that denitrification occurred predominantly in the upwelling areas. Analyses of microbial communities in the hyporheic zone showed that the new genera, species, and isotope data were associated with the hydrological uniqueness of the hyporheic zones. The 16S rRNA sequences were determined and phylogenetic analysis revealed that the DNB communities distributed and gathered the genus Comamonas denitrificans within the mixing patterns of the hyporheic zones and that the relative scarcity of these microbes in these zones was caused by the lack of appropriate substrates. The delineation of the surface water–groundwater mixing zone was quantitatively determined by systematically combining the hydrological and heat transfer analyses and by comparing denitrifying bacteria communities and N isotope data. This study showed that pyrosequencing and isotopic approaches are useful for evaluating the transformation processes of nitrate at the upwelling and downwelling points of a hyporheic zone. [ABSTRACT FROM AUTHOR]

An integrated study based on hydrogeochemical, microbiological and dual isotopic approaches for nitrate and sulfate was conducted to elucidate sources and biogeochemical reactions governing groundwater contaminants in different seasons and under different land use in a basin of Korea. The land use in the study area is comprised of forests (58.0%), vegetable fields (27.6%), rice paddy fields (11.4%) and others (3.0%). The concentrations of NO 3 –N and SO 4 2 − in groundwater in vegetable fields were highest with 4.2–15.2 mg L − 1 and 1.6–19.7 mg L − 1 respectively, whereas under paddy fields NO 3 –N concentrations ranged from 0 to 10.7 mg L − 1 and sulfate concentrations were ~ 15 mg L − 1 . Groundwater with high NO 3 –N concentrations of > 10 mg L − 1 had δ 15 N–NO 3 − values ranging from 5.2 to 5.9‰ and δ 18 O values of nitrate between 2.7 and 4.6‰ suggesting that the nitrate was mineralized from soil organic matter that was amended by fertilizer additions. Elevated concentrations of SO 4 2 − with δ 34 S–SO 4 2 − values between 1 and 6‰ in aquifers in vegetable fields indicated that a mixture of sulfate from atmospheric deposition, mineralization of soil organic matter and from synthetic fertilizers is the source of groundwater sulfate. Elevated δ 18 O–NO 3 − and δ 18 O–SO 4 2 − values in samples collected from the paddy fields indicated that denitrification and bacterial sulfate reduction are actively occurring removing sulfate and nitrate from the groundwater. This was supported by high occurrences of denitrifying and sulfate reducing bacteria in groundwater of the paddy fields as evidenced by 16S rRNA pyrosequencing analysis. This study shows that dual isotope techniques combined with microbial data can be a powerful tool for identification of sources and microbial processes affecting NO 3 − and SO 4 2 − in groundwater in areas with intensive agricultural land use. [ABSTRACT FROM AUTHOR]