Distribution of anammox bacteria in a free-water-surface constructed wetland with wild rice (Zizania latifolia)

•Anammox activity varied spatially in a free-water-surface constructed wetland.•The upper soil layer had higher anammox activity than deeper layers.•Highest anammox activity was at a submerged, vegetated site close to the inflow.•Anammox rRNA genes show similarity to anammox from fresh-water environments.

We investigated the distribution of anaerobic ammonium oxidation (anammox) bacteria under heterogeneous environmental conditions in a free-water-surface constructed wetland planted with wild rice (Zizania latifolia). The wetland receives effluent from a domestic wastewater treatment facility. Potential anammox activity was estimated using a nitrogen isotope pairing technique: activity was highest in the upper soil layer (0–2 cm). We also surveyed the horizontal distribution of potential anammox activity at seven sites over the wetland under different environmental conditions, along with copy numbers of anammox genes, which were estimated using quantitative PCR analysis. Three submerged sites showed substantial potential anammox activity and relatively high anammox copy numbers. The highest activity and highest copy number were 50.3 nmol N2 g-dry matter−1 h−1 and 1.42 × 1013 copy g-DNA−1, respectively, observed at a site that was submerged, vegetated, and immediately adjacent to the inflow from the domestic wastewater treatment facility. Phylogenetic analysis of anammox bacterial 16S rRNA genes revealed anammox bacteria with high similarity to uncultured anammox bacteria from freshwater environments such as paddy fields, and close to species in genera including “Candidatus Brocadia” and “Ca. Jettenia”. The highest contribution of potential anammox activity to potential total N2 gas production was 3.1%. This study provides new and useful information about the distribution of anammox in a constructed wetland treating actual wastewater.

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