Bacillus subtilis CECT30572 for biological fertilizer stabilization: an alternative to synthetic nitrification inhibitors
- 1Chemical, Environmental and Bioprocess Engineering Group, Institute for Research and Innovation in Engineering (I4), University of León
- 2Centro de Tecnologías Agroambientales (CTA) Fertiberia - Edificio Centro de Investigación, Tecnología e Innovación de la Universidad de Sevilla (CITIUS) (Centro de Investigación, Tecnología e Innovación) 1
Abstract
Introduction: Modern intensive agriculture faces the critical challenge of reduced nitrogen use efficiency and severe environmental impacts associated with nitrate (NO ₃ ⁻ ) leaching. Although chemical nitrification inhibitors such as 3,4-dimethylpyrazole phosphate (DMPP) are widely used to reduce nitrogen losses, there is growing interest in sustainable, bio-based alternatives. This study investigates Bacillus subtilis CECT30572, the active biological component of the commercial nitrogen stabilizer NSAFE ® , as a microbial strategy to reduce nitrate leaching.
Methods: Hybrid whole-genome sequencing was performed to characterize genes associated with nitrogen metabolism. In vitro assays evaluated nitrate depletion, and soil column leaching experiments were conducted under contrasting soil types using different fertilizer formulations.
Results: Hybrid whole-genome sequencing revealed a conserved nasABCDEF cluster and regulatory elements ( tnrA, glnR, codY, gltC ) associated with nitrogen assimilation. The genome also encodes the respiratory narGHJI operon for nitrate reduction to nitrite (NO ₂ > ⁻ ), while lacking canonical genes for complete denitrification ( nirK/S, norBC, nosZ ), indicating a non-denitrifying nitrogen metabolism. In vitro assays showed substantial nitrate removal, with a net depletion of 455 mg L ⁻¹ NO ₃ ⁻¹ . The nitrate removed from the medium was not recovered predominantly as extracellular or intracellular-associated NH ₄ ⁺ . Although its specific fate cannot be determined from the present measurements, we hypothesize that it could have been assimilated with the bacteria in the form of amino acids. Soil column leaching experiments under contrasting soil types showed that coating or mixed fertilizers (granular NP, granular N, UAN) with CECT30572 significantly reduced cumulative NO ₃ ⁻ leaching, with reductions of a magnitude similar to those observed with DMPP under several experimental conditions.
Discussion: Overall, these results indicate that the nitrate removal observed for B. subtilis CECT30572 under in vitro conditions may contribute to the reduction in nitrate losses observed in soil columns, supporting its potential as a bio-based strategy for reducing nitrate leaching.
Related
- The nitrification inhibitor DMPP preferentially suppresses ammonia-oxidising bacteria and reduces nitric oxide emissions in an archaeal-dominated grassland soil.
- Structure-activity relationship study in soil identifies an effective broad-spectrum inhibitor of archaeal nitrification
- The Influence of Planting Different Green Manures in Apple Orchards on Soil Fertilizer and Apple Quality in the Loess Plateau
- Organic fertilizer amendments reshape soil microbial communities and metabolic functions in sweet cherry production
- Agronomic Performance and Elemental Uptake of a Potential Fertilizer Derived from Microwave-Treated Animal By-Products in Lepidium sativum and Lactuca sativa
BibTeX
@article{mazuecosaguilera2026bacillus,
title = {{Bacillus subtilis CECT30572 for biological fertilizer stabilization: an alternative to synthetic nitrification inhibitors}},
author = {Ismael Mazuecos-Aguilera and Pablo Fernández-Carbajo and Ana M. Laureano-Marín and Javier Brañas and Fernando González-Andrés},
journal = {Frontiers in Plant Science},
year = {2026},
month = oct,
doi = {10.3389/fpls.2026.1936312},
url = {https://www.frontiersin.org/articles/10.3389/fpls.2026.1936312/full},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Bacillus subtilis CECT30572 for biological fertilizer stabilization: an alternative to synthetic nitrification inhibitors
AU - Ismael Mazuecos-Aguilera
AU - Pablo Fernández-Carbajo
AU - Ana M. Laureano-Marín
AU - Javier Brañas
AU - Fernando González-Andrés
PY - 2026
DA - 2026/10/29
JO - Frontiers in Plant Science
DO - 10.3389/fpls.2026.1936312
UR - https://www.frontiersin.org/articles/10.3389/fpls.2026.1936312/full
AB - Introduction: Modern intensive agriculture faces the critical challenge of reduced nitrogen use efficiency and severe environmental impacts associated with nitrate (NO ₃ ⁻ ) leaching. Although chemical nitrification inhibitors such as 3,4-dimethylpyrazole phosphate (DMPP) are widely used to reduce nitrogen losses, there is growing interest in sustainable, bio-based alternatives. This study investigates Bacillus subtilis CECT30572, the active biological component of the commercial nitrogen stabilizer NSAFE ® , as a microbial strategy to reduce nitrate leaching. Methods: Hybrid whole-genome sequencing was performed to characterize genes associated with nitrogen metabolism. In vitro assays evaluated nitrate depletion, and soil column leaching experiments were conducted under contrasting soil types using different fertilizer formulations. Results: Hybrid whole-genome sequencing revealed a conserved nasABCDEF cluster and regulatory elements ( tnrA, glnR, codY, gltC ) associated with nitrogen assimilation. The genome also encodes the respiratory narGHJI operon for nitrate reduction to nitrite (NO ₂ > ⁻ ), while lacking canonical genes for complete denitrification ( nirK/S, norBC, nosZ ), indicating a non-denitrifying nitrogen metabolism. In vitro assays showed substantial nitrate removal, with a net depletion of 455 mg L ⁻¹ NO ₃ ⁻¹ . The nitrate removed from the medium was not recovered predominantly as extracellular or intracellular-associated NH ₄ ⁺ . Although its specific fate cannot be determined from the present measurements, we hypothesize that it could have been assimilated with the bacteria in the form of amino acids. Soil column leaching experiments under contrasting soil types showed that coating or mixed fertilizers (granular NP, granular N, UAN) with CECT30572 significantly reduced cumulative NO ₃ ⁻ leaching, with reductions of a magnitude similar to those observed with DMPP under several experimental conditions. Discussion: Overall, these results indicate that the nitrate removal observed for B. subtilis CECT30572 under in vitro conditions may contribute to the reduction in nitrate losses observed in soil columns, supporting its potential as a bio-based strategy for reducing nitrate leaching.
SN - 1664-462X
ER -