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The Florida Everglades is historically limited in phosphorus; however, runoff from the Everglades Agricultural Area has resulted in a gradient in phosphorus concentrations running into the interior of the northern Everglades. Phosphorus enrichment resulted in a shift from P- to N-limitation along the gradient, such that a detailed understanding of the impacts of P-enrichment on the nitrogen cycle is needed for a complete understanding of nutrient impacts and restoration ecology in the Everglades. This study aimed to investigate nitrogen fixation rates and the distribution of genes encoding dinitrogenase reductase (nifH) in soils along the nutrient gradient in Water Conservation Area 2-A, including a site previously exposed to high levels of nutrients (F1), a transition site (F4), and a site unimpacted by nutrient additions (U3). At the 0-2 cm depths, the highest acetylene reduction rates were observed in F4 (33 µmol C2H4/(g soil•h)), followed by F1 (15 µmol C2H4/(g soils•h) and U3 (1.4 µmol C2H4/(g soils•h). These general trends were corroborated by 15N uptake studies. These rates are in good agreement with the abundance of nifH measured by a real-time PCR: F4, 7109; F1, 5109; U3, 1 109 (in gram soil). The community analysis of nitrogen-fixing microbes using nifH clone libraries indicated that a broad range of phylogenetic groups, including representatives of the Alpha-, Beta-, Gamma-, and Epsilonproteobacteria, Firmicutes (primarily Clostridium sp.), and other anaerobes. Significantly, sequences related to methanogenic Archaea were detected in all sites, with the greatest proportion observed in F4 (~32%) compared with other sites (F4, 9%; U3, 12%). Hydrogen, a common substrate for methanogenesis, was demonstrated to be a favorable substrate for anaerobic nitrogen fixation. These observations indicate that hydrogen is an important factor regulating both nitrogen-fixation rates and methanogenesis in Everglades. |