Two methods are commonly employed to quantify rates of N₂-fixation in surface waters. Acetylene reduction assays (ARA) are an indirect measure of N₂-fixation, relying on the conversion of acetylene to ethylene at a known mole ratio with respect to dissolved nitrogen gas. An alternative method, membrane-inlet-mass-spectrometry (MIMS), can directly quantify dissolved N₂ gas concentrations in water.Here, surface sample grabs (0.5m depth) were collected at 6 sites in Sandusky Bay on 4 dates (N=24) and bioassays were performed using both techniques. Significant levels of N2-fixation were detected with ARA in 83% of samples, whereas net N2-fixation was not detected in any samples (0%) by analysis via MIMS. In contrasts, significant levels of N2-production were observed in 40% of the samples. N2-production is not possible to detect with ARA and would have been missed had MIMS not been employed. Notably, 36% of the samples yielded significant rates of N2-fixation via ARA and also net N2-production via MIMS. In order for analysis via MIMS to show significant increases in concentrations of dissolved N2, the rates of N2-production must exceed the rates of N2-fixation in those samples. It is well documented that members of the cyanobacterial order Nostocales produce specialized cells (heterocyst) that can fix nitrogen, converting inert gaseous N2 into ammonium ion (NH4+). Thus, detecting N2-fixation in Sandusky Bay is not atypical. However, N2-production is known to occur via two pathways within the microbial nitrogen cycle (i.e., anaerobic ammonium oxidation and denitrification). Currently, neither of these N2-production pathways are known to be associated with oxygenated surface waters, but rather anoxic lake sediments (denitrification) or groundwater (anaerobic ammonium oxidation). Additional research is needed to resolve the net N2-production observed by the methods employed in this study.

Historic coal mines are one of the major generators of Acid Mine Drainage (AMD) in the United States. Past coal mining activities in Huff Run, Ohio generated large mine spoils and oxidation of these mine spoils led to a serious AMD generation in the Huff Run watershed. A remediation project conducted in the Huff Run watershed costing over $590,000 failed to resolve AMD generation. The persistent AMD generation could be associated with the possible formation, transportation and oxidation of colloidal pyrite from mine spoils. To investigate the transport of colloidal pyrite through unsaturated soil in Huff Run subwatershed HR#25, eight lysimeters were installed and pore water samples were collected during summer of 2018. Initial studies have indicated the presence of colloidal pyrites in the porewater. The scanning electron microscopy-energy dispersive spectrometer (SEM-EDS) analysis of the colloidal particles isolated from pore water samples revealed the presence of iron (Fe), sulfur (S) and oxygen (O) bearing phases in the colloids which are consistent with pyrite (FeS2) and the oxidation product Fe oxides. The x-ray diffraction (XRD) analysis of the colloidal samples confirmed the presence of multiple phases of sulfide bearing minerals (FeS2, FeAsS2, CuFeS2). Both SEM-EDS and XRD detected the presence of Fe oxides associated with colloidal pyrite which can have a significant environmental impact in terms of colloidal pyrite oxidation, AMD generation and trace metals adsorption.