Vertical stratification of microbial communities and isotope geochemistry tie groundwater denitrification to sampling location within a nitrate-contaminated aquifer

Nitrate pollution is a major threat to groundwater quality in agricultural areas. Natural attenuation of nitrate in contaminated aquifers is mediated by denitrifying microbial populations in anoxic environments. Vertical distribution of denitrifying microbial communities in aquifers is greatly influenced by groundwater redox conditions, local hydrogeological parameters, and seasonal variability in groundwater flow and recharge. In this study, we investigated groundwater geochemistry and the composition of bacterial and archaeal communities with increasing depth in a shallow nitrate-contaminated aquifer in British Columbia, Canada. High-resolution passive diffusion sampling was conducted to collect groundwater at 10-cm intervals from 4 to 20 m below ground surface (mbgs) in the aquifer. Geochemical analyses of major ions indicated a general shift in the groundwater chemistry below 16 mbgs including decreasing chloride concentrations that suggest two-end member mixing of shallow and deep groundwater with different chemistries. A redoxcline was further observed within a 2 m transition zone at 18-20 mbgs characterized by sharp declines in nitrate concentrations and increases in sulfate and total inorganic carbon. Excursions in delta N-15-NO3- and delta O-18-NO3- in the same depth interval are consistent with denitrification, and a concomitant decrease in delta S-34-SO42- suggested that denitrification was coupled to sulfide or sulfur oxidation. Microbial communities within this depth interval were significantly dissimilar to those above and below, featuring putative lithotrophic denitrifying bacteria belonging to the genera Sulfurifustis, Sulfuritalea and Sulfuricella. These lineages were detected in greatest abundance at 19 mbgs while the abundances of putative heterotrophic sulfate-reducing bacteria belonging to the genus Desulfosporosinus were greatest at 20 mbgs. In addition to help distinguish denitrification from mixing-induced changes in groundwater chemistry, the above observed vertical stratification of the microbial key players connects nitrate removal to the locations of the aquifer sampled.