Combined use of δ(13)C, δ(15)N, and δ(34)S tracers to study anaerobic bacterial processes in groundwater flow systems.

We present an approach for determining the major anaerobic bacterial processes in aquifers, using the combined stable isotope ratios of major elements (C, N, and S) as net recorders of the biogeochemical reactions. The Kumamoto groundwater is constituted of two major flow systems, A-A' and B-B', within 10(3) km-scale area. Previous study [Hosono, T., Tokunaga, T., Kagabu, M., Nakata, H., Orishikida, T., Lin, I-T., Shimada, J., 2013. The use of δ(15)N and δ(18)O tracers with an understanding of groundwater flow dynamics for evaluating the origins and attenuation mechanisms of nitrate pollution. Water Res. 47, 2661-2675.] investigated the nitrate sources and extent of denitrification using [Formula: see text] and [Formula: see text] tracers. In the present study, we studied a type of denitrification (heterogenic vs. autotrophic) and occurrence of sequential anaerobic processes along the flow systems with newly measured δ(13)CDIC and [Formula: see text] . In A-A' flow system, C, N, and S isotopic compositions did not change along the flow direction. However, in B-B' flow system significant sulfate reduction (with a maximum [Formula: see text] enrichment of +55‰) occurred along with denitrification (with a maximum [Formula: see text] enrichment of +38‰) as the groundwater flowed down-gradient. Depletions in [Formula: see text] (-8‰ maximum) were found only sporadically. These observations imply that autotrophic denitrification could occur in very limited parts of the study area. Moreover, the occurrence of methanogenic reactions was suggested by the enriched δ(13)CDIC signature (+8‰ maximum) at a denitrification hotspot. By characterizing C, N, and S isotope compositional changes in wide redox range (from aerobic oxidation of organic carbon, through denitrification, to sulfate reduction, until methanogenesis), we could develop the model of C, N, and S isotopic evolutional patterns under different redox scenarios. Proposed model is useful in obtaining a comprehensive understanding of the major anaerobic bacterial processes in aquifer systems, including distinguishing between heterotrophic and autotrophic denitrification.