Coupled molecular and isotopic evidence for denitrifier controls over terrestrial nitrogen availability

Denitrification removes ecologically available nitrogen (N) from the biosphere and influences both the pace and magnitude of global climate change. Disagreements exist over the degree to which this microbial process influences N-availability patterns across Earth's ecosystems. We combine natural stable isotope methods with qPCR to investigate how denitrifier gene abundance is related to variations in nitrate (NO₃^-) pool sizes across diverse terrestrial biomes and conditions. We analyze NO₃^- isotope composition (¹⁵N/¹⁴N, ¹⁸O/¹⁶O) and denitrifier gene nirS in 52 soil samples from different California ecosystems, spanning desert, chaparral, oak-woodland/savanna and forest. δ¹⁵N-NO₃^- correlates positively with δ¹⁸O-NO₃^- (P⩽0.03) and nirS abundance (P=0.00002) across sites, revealing the widespread importance of isotopic discrimination by soil denitrifiers. Furthermore, NO₃^- concentrations correlate negatively to nirS (P=0.002) and δ¹⁵N-NO₃^- (P=0.003) across sites. We also observe these spatial relationships in short-term (7-day), in situ soil-incubation experiments; NO₃^--depletion strongly corresponds with increased nirS, nirS/16 rRNA, and enrichment of heavy NO₃^- isotopes over time. Overall, these findings suggest that microbial denitrification can consume plant-available NO₃^- to low levels at multiple time scales, contributing to N-limitation patterns across sites, particularly in moist, carbon-rich soils. Furthermore, our study provides a new approach for understanding the relationships between microbial gene abundance and terrestrial ecosystem functioning.