Little Lagoon is a shallow coastal lagoon that is tidally connected to the Gulf of Mexico but has no riverine inputs. The water in the lagoon is replenished solely from precipitation and groundwater inputs primarily on the East end (Su et al. 2012). Because of the rapid development in Baldwin County, a large amount of NO3- enters the Little Lagoon system through SGD (Murgulet & Tick 2008). In this region, there can be rapid changes in the depth to groundwater (Fig. 4.1 inset) and episodic SGD inputs to the lagoon (Su et al.2013). Within the lagoon, three sites were selected (East, Mouth, and West) to represent the gradient that exists across the lagoon from the input of groundwater. Sites were sampled on a near-monthly basis from February 2012 to February 2013.

Potential denitrification and N2 fixation from slurry assays

The top 50 mm of sediment at each site were collected in duplicate with a large core (95 mm ID) and homogenized. Potential denitrification rates were measured following the acetylene inhibition technique (Sørensen 1978) but may underestimate denitrification supported by coupled nitrification-denitrification, as this method inhibits nitrification. To triplicate serum vials, approximately 20 g of sediments and filtered (0.7 micron) site water were added at various treatments (control and N addition (100 μM and 500 μM KNO3 -). Samples were sealed with a butyl rubber stopper, capped and flushed with N2 gas for 10 minutes. After the addition of C2H2 (10% v/v) and a 1-hour incubation, headspace gas samples were injected into evacuated 12 ml Exetainer vials and N2O production was quantified with a Shimadzu GC-2014 with an electron capture detector (GC-ECD) within 24 hours.

Potential N2 fixation rates were measured as ethylene (C2H4) production from acetylene (C2H2) reduction (Welsh et al. 1996) in triplicate from slurry assays containing 20 g of homogenized sediment and filtered (0.7 micron) site water. Rates of N2 fixation by sulfate reducing bacteria (SRB) were determined after the addition of sodium molybdate as a specific inhibitor of the sulfate reduction process (Hardy et al. 1973, Capone 1993). After C2H4 analysis on a Shimadzu gas chromatograph (GC-2014) with flame ionization detection (GC-FID), production rates of C2H4 were converted to potential N2 fixation rates using a C2H2:N2 reduction ratio of 3:1 (Capone 1993).

Additional methodology can be found in:

Bernard, Rebecca & Mortazavi, Behzad & A. Kleinhuizen, Alice. (2015). Dissimilatory nitrate reduction to ammonium (DNRA) seasonally dominates NO3− reduction pathways in an anthropogenically impacted sub-tropical coastal lagoon. Biogeochemistry. 125. 47-64. 10.1007/s10533-015-0111-6.