Stimulating Denitrification of Agricultural Drainage Using Elemental Sulfur: Denitrification Rates and Microbial Community Structure

Agricultural headwater streams are a major pathway for the export of nitrate (NO3−) into the environment. These streams may also be well-suited for removing NO3−, via microbial denitrification, thereby preventing export to downstream ecosystems. Since denitrification is often limited by the lack of readily degradable electron donor, we explored the potential for amending agricultural streams with elemental sulfur granules, which develop sulfur-oxidizing, denitrifying biofilms. Our experimental system was a stream mesocosm lined with elemental sulfur granules. With a nitrogen loading rate of 240 mgN m−2 d−1, denitrification fluxes were as high as 228 mgN m−2 d−1, which is 5 to 35 times higher than reported previously for agricultural drainage streams. Over 64 weeks of the study, NO3− removal efficiencies decreased over time, concurrent with accumulation of biofilm biomass on the benthic bed surface, suggesting that accumulation of biomass or fine sediments can limit removal efficiencies. Sulfate (SO42−) concentrations exceeded the stoichiometric amount for sulfur-based denitrification alone indicating oxygen reduction by sulfur-oxidizing bacteria was also occurring. Molecular studies demonstrated that the sulfur amendment selected for Thiobacillus species, and that no special inoculum was required for establishing a sulfur-based autotrophic denitrifying community. A mathematical model of the stream mesocosm NO3− removal was developed to describe the biological reactions, accurately capturing trends in bulk liquid NO3− concentrations when a high diffusive exchange coefficient was used. Biological activity was likely limited to the top few millimeters of the sulfur bed where advective exchange is likely to be significant. Results suggest sulfur amendment is effective for enhancing denitrification in agricultural streams. However, placement of sulfur deeper in the streambed may minimize SO42− formation and reduce the potential for incomplete denitrification to N2O.