Two nitrification inhibitors showed distinct effects in a soil microbe, suggesting they may interfere with ammonia oxidation through different cellular routes. In laboratory cultures of the archaeon Nitrososphaera viennensis, DMPP was associated with an energy-limitation response, while ATU was linked to broader repression of gene activity and energy-related pathways.
The study examined how DMPP and ATU affect N. viennensis and whether their effects could be relieved by adding ammonium, urea or copper. The researchers combined oxygen-consumption tests with growth measurements and multiomics, comparing changes in RNA transcripts and proteins.
The response changed with the substrate
The first clue came from microrespiration tests, which tracked oxygen use as an indicator of metabolic activity. DMPP was associated with 59.6% inhibition when ammonium was supplied, but with 24.1% inhibition when hydroxylamine was used instead. ATU was effective only in the presence of ammonium.
This substrate-dependent pattern made the two compounds look different at the level of the organism’s metabolism. It also led to relief experiments designed to test whether adding more substrate could change the response. The pattern did not by itself establish a molecular binding site or inhibition mechanism.
DMPP moved through the cell’s response system first
The researchers followed DMPP-treated cultures over 12 hours, taking samples after 0.5, 1, 3, 6 and 12 hours. Changes in RNA transcripts could be separated from controls as early as 0.5 hours, while differences in the measured protein profile became apparent only after at least six hours. Sequencing averaged 26.45 million paired-end reads per sample, and the protein analysis covered 1,032 of 3,187 predicted proteins.
In a separate comparison, cultures received no inhibitor, 2 mM DMPP or 0.5 mM ATU. Each condition had eight cultures, with five used for molecular measurements and three for growth monitoring. A multivariate analysis using selected genes or proteins separated untreated cultures from inhibitor-treated cultures and also distinguished ATU from DMPP.
One response looked like energy shortage, the other like a deeper pause
The DMPP-treated cells showed an energy-limitation response. They increased expression of genes involved in urease production, along with a nearby transporter for urea. The ATU-treated cells showed the more severe physiological response, with widespread transcriptional repression and lower activity in pathways tied to energy metabolism.
The authors interpreted the contrast as consistent with different inhibition modes. They described DMPP as strongly suggestive of competitive inhibition and ATU as consistent with noncompetitive inhibition and a quiescence-like cellular state. Those interpretations remain inferential because the study did not directly measure inhibitor binding to the ammonia monooxygenase complex or carry out direct inhibition-kinetic tests.
Extra copper did not restore activity
Adding copper or other trace elements did not relieve inhibition in either the ATU or DMPP cultures. That made a primary mechanism based on chelating free copper less likely under the tested conditions, although the study did not completely exclude every possible interaction between DMPP and copper.
The substrate tests produced a sharper split. Adding 2.5 mM urea or 5 mM ammonium was followed by relief of stress in DMPP-treated cultures, while 1 mM urea was not enough. The same additions did not relieve the stress associated with ATU.
A result from one cultured strain
All of the experiments used N. viennensis EN76, originally from garden soil in Vienna, grown aerobically in dark, shaking cultures at 80 rpm, 42 °C and pH 7.5 in 10 mM medium supplemented with 2 mM ammonium chloride. The work therefore provides mechanistic evidence from one laboratory strain under controlled conditions, rather than a field test across soils or other nitrifying organisms.
The tested inhibitor concentrations should not be treated as direct equivalents of exposure levels in soil. The proposed mechanisms also remain open to further testing, including comparisons with other nitrifiers and direct biochemical studies of the ammonia monooxygenase complex.
Paper data and sources
Original title: Common Nitrification Inhibitors Exhibit Distinct Mechanisms on the Ammonia-Oxidizing Archaeon Nitrososphaera viennensis
Authors: Dalkidis D, Malits A, Kerou M et al.
Journal/Repository: Environmental science & technology
Status: Peer-reviewed
First online: 2026-10-19
DOI: Not available
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