posted on 2024-01-25, 03:33authored byLinqi Zhang, Huicheng Zhao, Shuping Qin, Chunsheng Hu, Yanjun Shen, Baoyuan Qu, Yang Bai, Binbin Liu
The
heavy use of nitrogen fertilizer in intensive agricultural
areas often leads to nitrate accumulation in subsurface soil and nitrate
contamination in groundwater, which poses a serious risk to public
health. Denitrifying microorganisms in the subsoil convert nitrate
to gaseous forms of nitrogen, thereby mitigating the leaching of nitrate
into groundwater. Here, we investigated denitrifying microorganisms
in the deep vadose zone of a typical intensive agricultural area in
China through microcosm enrichment, genome-resolved metagenomic analysis,
and denitrifying bacteria isolation. A total of 1000 metagenome-assembled
genomes (MAGs) were reconstructed, resulting in 98 high-quality, dereplicated
MAGs that contained denitrification genes. Among them, 32 MAGs could
not be taxonomically classified at the genus or species level, indicating
that a broader spectrum of taxonomic groups is involved in subsoil
denitrification than previously recognized. A denitrifier isolate
library was constructed by using a strategy combining high-throughput
and conventional cultivation techniques. Assessment of the denitrification
characteristics of both the MAGs and isolates demonstrated the dominance
of truncated denitrification. Functional screening revealed the highest
denitrification activity in two complete denitrifiers belonging to
the genus Pseudomonas. These findings greatly expand
the current knowledge of the composition and function of denitrifying
microorganisms in subsoils. The constructed isolate library provided
the first pool of subsoil-denitrifying microorganisms that could facilitate
the development of microbe-based technologies for nitrate attenuation
in groundwater.