Denitrification Activity and Relevant Bacteria Revealed by Nitrite Reductase Gene Fragments in Soil of Temperate Mixed Forest

Abstract

Denitrification activity and bacterial community constituents were investigated in both well-drained and poorly drained soils of a temperate forest in central Japan by ¹⁵N tracer experiments and a cloning-sequencing approach. Denitrification activity was much higher in wet soil than in dry soil, based on ¹⁵N¹⁵N (³⁰N₂) and ¹⁵N¹⁵NO (⁴⁶N₂O) production. Labeled nitrate (¹⁵NO₃^-) was immediately reduced to ³⁰N₂ in wet soil, whereas it was only reduced to ⁴⁶N₂O in dry soil. Thus, the wet soil at the lower end of the catchment is a functional site for the scavenging for NO₃^- and N₂O. Nitrite reductase gene (nirK and nirS) fragments from these soils were PCR amplified, cloned, and sequenced. Both nirK and nirS fragments were detected in the wet soil, whereas only nirK fragments were detected in the dry soil. All the nirK and nirS clones showed less than 90% similarity to known clones. Numerous operational taxonomic units for nirK and nirS were found in the wet soil. Considerable diversification within the largest clade on the nirK phylogenetic tree, which contained no known sequence, was observed in wet soil. Thus, a wet soil environment can provide both the habitat and conditions for the expression of denitrification activity.