Influence of Nitrate and Phosphorus Loading on Denitrifying Enzyme Activity in Everglades Wetland Soils

There has been recent concern about the impact of increased nutrient loading on the northern Everglades ecosystem. We investigated the spatial and temporal distribution of denitrifying enzyme activity (DEA) along a P-enrichment gradient in the Water Conservation Area 2A (WCA-2A) and determined the effects of added P and NO., on DEA. The DEA in soil and detritus layers was measured under anaerobic conditions four times during 2 yr, using the acetylene blockage technique. The DEA ranged from 0.004 to 7.75 mg NjO-N kg" h~'. Highest rates of DEA were found in the detritus and surface (0-10 cm) soils, and rates decreased exponentially with increasing distance from the surface-water inflow point, where nutrients are loaded to the wetland. Nitrate was found to be limiting, while the addition of P had no effect on the distribution of DEA in these soils. There was a seasonal effect on DEA, with higher activity observed during the summer when temperatures and hydraulic and nutrient loading were highest. Soils from outside the impacted zone demonstrated denitrifying potentials, within 10 h when spiked with inflow concentrations of NOf, similar to DEA of soils from within the impacted zone. This suggests that soils from outside the impacted zone can increase denitrification rates when exposed to higher NOf concentrations in a relatively short time. Agricultural drainage water discharge, and consequent NOr loading, has created a zone of elevated DEA proximal to the S-lOCsnrface-water inflow point in WCA-2A. N REDUCTION is the major N removal mechanism in wetlands. Among NO3~ reduction processes, denitrification is the dominant NO3~ removal process. Denitrification is a microbially mediated process whereby facultative anaerobic bacteria use NO3~ (or NOr) in the absence of O2 as the terminal electron acceptor during the oxidation of organic C (microbial respiration), resulting in the production of gaseous end products, N2O and N2. The denitrification enzyme assay is used as a means to eliminate all other regulating factors of denitrification in order to quantify the amount of active denitrifying enzymes present in soil (Smith and Tiedje, 1979; Smith and Parsons, 1985; Groffman, 1987; Schipper et al., 1993). The enzyme assay is the shortterm (2 h) rate of N2O production and is indicative of the size and activity of the denitrifying enzyme pool present in soil. The assay reflects the immediate biological effect of changes in redox conditions attributed to changes in soil O2 levels (Martin et al., 1988). Several studies of DEA have focused on upland soils Univ. of Florida, Wetland Biogeochemistry Lab., 106 Newell Hall, P.O. Box 110510, Gainesville, FL 32611. Florida Agricultural Experiment Station Journal Series no. R-06680. Received 17 Dec. 1998. * Corresponding author ([email protected]). Published in Soil Sci. Soc. Am. J. 63:1945-1954 (1999). (Smith and Tiedje, 1979; Groffman, 1987; Parsons et al., 1991), with the goal of more recent studies focused on correlating denitrification potential at the ecosystem scale to easily measurable soil parameters. These studies have reported rates of N2O production in upland soils ranging from 0.006 to 7.14 mg N kg" h". The results of such research could be used to quantify the contribution of soils to global atmospheric N2O levels. Several problems exist with the use of DEA soil measurements in extrapolating to landscape scale denitrification rates in upland soils. The microorganisms responsible for the production of enzymes are facultative anaerobes, which possess separate enzyme systems capable of using either O2 or NO} as terminal electron acceptors. The NO3~ reducing enzyme systems are primarily inactive in the presence of O2 and active in enzyme production only during ephemeral anoxic events (e.g., rainfall events; Sexstone et al., 1985; Burton and Beauchamp, 1985). Further, the presence of O2 appears to repress or deactivate enzymes already present in soil (Martin et al., 1988). Secondly, the distribution of organic matter in upland systems is patchy, which presents additional problems in assessing the spatial distribution of DEA. Hotspots of organic matter provide both simple C compounds for the maintenance of large, microbial populations and anoxic microenvironments that lead to NO3~ consumption as the terminal electron acceptor (Parkin, 1987; Christensen et al., 1990a, 1990b). The patchy distribution of active enzyme sites in the landscape leads to logarithmic frequency distributions of enzyme activity measured in the field (Parkin, 1987). Finally, NO3~ is rarely the limiting factor for denitrification in upland ecosystems, as evidenced by the widespread problem of groundwater contamination of NO3~, and consequently is a poor indicator of denitrification potential. These factors in combination confound efforts to consistently correlate easily measured soil parameters (total C, water content, NO3~ concentration, and microbial biomass) with DEA to produce meaningful estimates of denitrification in the landscape. (Parsons et al., 1991; Velthof, 1996). There exist several important differences between mineral, upland soils and organic-rich, wetland soils that permit the reliable use of DEA on a landscape scale in order to investigate the source and effect of NO-T loading in wetlands. Wetland soils are often saturated most Abbreviations: ANOVA, analysis of variance; DEA, denitrifying enzyme activity; SFWMD, South Florida Water Management District; WCA-2A, Water Conservation Area 2A.