AME 66:283-293 (2012)  -  DOI: https://doi.org/10.3354/ame01574

ABSTRACT: Understanding the mechanistic controls of microbial denitrification is of central importance to both environmental microbiology and ecosystem ecology. Loss of nitrate (NO₃⁻) is often attributed to carbon-driven (heterotrophic) denitrification. However, denitrification can also be coupled to sulfur (S) oxidation by chemolithoautotrophic bacteria. In the present study, we used an in situ stable isotope (¹⁵NO₃⁻) tracer addition in combination with molecular approaches to understand the contribution of sulfur-oxidizing bacteria to the reduction of NO₃⁻ in a eutrophic lake. Samples were incubated across a total dissolved sulfide (H₂S) gradient (2 to 95 µM) between the lower epilimnion and the upper hypolimnion. Denitrification rates were low at the top of the chemocline (4.5 m) but increased in the deeper waters (5.0 and 5.5 m), where H₂S was abundant. Concomitant with increased denitrification at depths with high sulfide was the production of sulfate (SO₄²⁻), suggesting that the added NO₃⁻ was used to oxidize H₂S to SO₄²⁻. Alternative nitrate removal pathways, including dissimilatory nitrate reduction to ammonium (DNRA) and anaerobic ammonium oxidation (anammox), did not systematically change with depth and accounted for 1 to 15% of the overall nitrate loss. Quantitative PCR revealed that bacteria of the Sulfurimonas genus that are known denitrifiers increased in abundance in response to NO₃⁻ addition in the treatments with higher H₂S. Stoichiometric estimates suggest that H₂S oxidation accounted for more than half of the denitrification at the depth with the highest sulfide concentration. The present study provides evidence that microbial coupling of S and nitrogen (N) cycling is likely to be important in eutrophic freshwater ecosystems.

KEY WORDS: Denitrification · Nitrate reduction · Sulfur oxidation · Sulfur-driven denitrification · Sulfurimonas denitrificans · Sulfide · Wintergreen Lake