| Summary: | Seagrass beds are important coastal habitats that are diminishing globally. Nitrogen, a key nutrient, often limits seagrass growth. Nitrogen fixation provides new, bioavailable nitrogen to the plants. This study explores its importance and factors controlling rates in sediments colonized by two dominant taxa in Northwest Florida, <i>Thalassia testudinum</i> and <i>Halodule wrightii</i>, compared to unvegetated sediments. We hypothesized that nitrogen fixation rates would be greater in seagrass colonized sediments, particularly during high growth periods. We expected to observe a positive relationship between rates and porewater sulfide concentrations because sulfate reducers were the dominant diazotrophs in similar studies. Rates were higher in vegetated areas. In <i>H. wrightii</i> beds, nitrogen fixation was driven by the decreased availability of porewater ammonium relative to phosphorus. In <i>T. testudinum</i> beds, rates were highest during winter. Organic matter may be a controlling factor in all substrate types albeit the exact mechanism driving nitrogen fixation differs slightly. During the summer and fall, nitrogen fixation provided between 1–15% of <i>T. testudinum</i> nitrogen demand. Annually, nitrogen fixation provided 4% and 1% of <i>T. testudinum</i> and <i>H. wrightii</i> nitrogen demand, respectively. Nitrogen fixation was an important source of nitrogen during periods of senescence and dormancy when organic matter content was high.
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