Nitrogen Cycling Dynamics in Constructed Wetlands Treating Agricultural Runoff
DOI:
https://doi.org/10.5281/zenodo.22687910Keywords:
constructed wetland, denitrification, nitrous oxide, agricultural runoff, nosZ, microbial community, nitrogen removalAbstract
Constructed wetlands are widely deployed to intercept nitrate-laden agricultural runoff before it reaches receiving
waterways, but their nitrogen removal performance is highly variable between sites and seasons, and the
microbial process balance underlying that variability, specifically the ratio of complete denitrification to the
incomplete pathway that releases the potent greenhouse gas nitrous oxide, is rarely characterised directly
alongside removal efficiency. We monitored nitrogen cycling in three free-water-surface constructed wetlands
receiving tile-drain agricultural runoff over a full annual cycle, measuring inlet-outlet nitrate mass balance, direct
nitrous oxide flux by static chamber, sediment denitrification potential by acetylene-block assay, and microbial
community composition and denitrification-gene abundance (nirS, nirK, nosZ clade I and II) by quantitative PCR
and 16S rRNA amplicon sequencing, across four seasonal sampling campaigns. Mean annual nitrate removal
efficiency was 58.4% across the three wetlands but varied more than twofold, from 41.2% to 76.8%, and removal
was significantly higher in summer than winter at every site (mean 71.3% versus 39.6%, paired t-test P<0.001),
tracking closely with sediment temperature and denitrification potential. Nitrous oxide flux, however, did not track
nitrate removal efficiency in the same direction: the wetland with the highest nitrate removal also showed the
highest per-unit-nitrogen-removed nitrous oxide emission factor, associated with a significantly lower nosZ clade II
to nirS gene ratio than the other two wetlands (Kruskal-Wallis P=0.008), consistent with a smaller relative capacity
to complete the final denitrification step reducing nitrous oxide to inert dinitrogen gas. Seasonal shifts in nosZ
clade II abundance, rather than total denitrifier abundance, best predicted the nitrous oxide emission factor across
sites and seasons (linear mixed model, R2=0.61). These results indicate that nitrate removal efficiency and
greenhouse gas performance are not simply two faces of the same denitrification process in constructed wetlands,
and that the community-level balance of denitrification gene clades, not just overall microbial activity, is an
important and underused design and management target for wetlands intended to deliver nitrogen removal without
a disproportionate nitrous oxide trade-off.
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