Characterization of Enrichment Cultures of Anammox, Nitrifying and Denitrifying Bacteria Obtained from a Cold, Heavily Nitrogen-Polluted Aquifer

Abstract

Anammox bacteria related to Candidatus Scalindua were recently discovered in a cold (7.5 °C) aquifer near sludge repositories containing solid wastes of uranium and processed polymetallic concentrate. Groundwater has a very high level of nitrate and ammonia pollution (up to 10 and 0.5 g/L, respectively) and a very low content of organic carbon (2.5 mg/L). To assess the potential for bioremediation of polluted groundwater in situ, enrichment cultures of anammox, nitrifying, and denitrifying bacteria were obtained and analyzed. Fed-batch enrichment of anammox bacteria was not successful. Stable removal of ammonium and nitrite (up to 100%) was achieved in a continuous-flow reactor packed with a nonwoven fabric at 15 °C, and enrichment in anammox bacteria was confirmed by FISH and qPCR assays. The relatively low total N removal efficiency (up to 55%) was due to nonstoichiometric nitrate buildup. This phenomenon can be explained by a shift in the metabolism of anammox bacteria towards the production of more nitrates and less N2 at low temperatures compared to the canonical stoichiometry. In addition, the too high an estimate of specific anammox activity suggests that N cycle microbial groups other than anammox bacteria may have contributed significantly to N removal. Stable nitrite production was observed in the denitrifying enrichment culture, while no “conventional” nitrifiers were found in the corresponding enrichment cultures. Xanthomonadaceae was a common taxon for all microbial communities, indicating its exclusive role in this ecosystem. This study opens up new knowledge about the metabolic capabilities of N cycle bacteria and potential approaches for sustainable bioremediation of heavily N-polluted cold ecosystems.