Inhibition profile of three biological nitrification inhibitors and their response to soil pH modification in two contrasting soils

Abstract

Up to 70% of the nitrogen (N) fertilizer applied to agricultural soils is lost through microbially mediated processes, such as nitrification. This can be counteracted by synthetic and biological compounds that inhibit nitrification. However, for many biological nitrification inhibitors (BNIs), the interaction with soil properties, nitrifier specificity, and effective concentrations are unclear. Here, we investigated three synthetic nitrification inhibitors (SNIs) (DCD, DMPP, and nitrapyrin) and three BNIs [methyl 3(4-hydroxyphenyl) propionate (MHPP), methyl 3(4-hydroxyphenyl) acrylate (MHPA), and limonene] in two agricultural soils differing in pH and nitrifier communities. The efficacies of SNIs and BNIs were resilient to short-term pH changes in the neutral pH soil, whereas the efficacy of some BNIs increased by neutralizing the alkaline soil. Among the BNIs, MHPA showed the highest inhibition and was, together with MHPP, identified as a putative AOB/comammox-selective inhibitor. Additionally, MHPA and limonene effectively inhibited nitrification at concentrations comparable to those used for DCD. Moreover, we identified the effective concentrations at which 50% and 80% of inhibition is observed (EC50 and EC80) for the BNIs, and similar EC80 values were observed in both soils. Overall, our results show that these BNIs could potentially serve as effective alternatives to SNIs currently used.

Keywords: EC50; agricultural soils; ammonia oxidation; biological nitrification inhibitors; pH.

Figure 1.

Soil microbial community composition in the AS and the CS. (A) 16S rRNA gene-based microbial community composition depicting the top taxa with ≥2% mean relative abundance at each site (B) amoA microbial community composition depicting the top amoA taxa with ≥5% mean relative abundance. Taxa with mean relative abundance below 2% (for 16S rRNA gene) or 5% (for amoA) are clustered as ‘other’. Samples collected in spring 2022 (n = 4, error bars represent standard error) (*P < .001). Absolute abundances of the amoA genes are shown in Supplementary Fig. 3.

Figure 2.

Abundance of ammonia oxidizers, total net nitrification potential activity, and AOA contribution to the net nitrification potential. (A) Absolute abundance of ammonia oxidizers in the AS and the CS. (B) Total net nitrification potential activity and AOA contribution to the net nitrification potential. Lower case letters depict significant differences within and between soils. Samples collected in spring 2022. Error bars represent the standard error in each case (n = 4, P < .05).

Figure 3.

Efficacy of SNIs and BNIs in the AS and CS. DMSO: dimethyl sulfoxide control; PA: phenylacetelyne. 1 mM PA, 4 µM Octyne, 100 µM ATU, 3.5 µM Nitrapyrin, 100 µM DCD, 10 µM DMPP, 200 µM MHPP, 200 µM MHPA, and 200 µM limonene. Samples collected in spring 2022. The median is depicted as the middle hinge in the boxplots. Upper and lower hinges represent the first and third quartile. The length of the whispers is determined by the largest and the smallest value in the dataset that are within 1.5 times the interquartile range. (n = 3, *P < .05, **P < .01, and ***P < .001, ns: not significant).

Figure 4.

Effect of soil pH manipulation on the efficacy of SNIs and BNIs in the (A) AS (pH 8.50), (B) CS (pH 6.12) PA: 1 mM Phenylacetelyne, 4 µM Octyne, 100 µM ATU, 3.5 µM Nitrapyrin, 100 µM DCD, 10 µM DMPP, 200 µM MHPP, 200 µM MHPA, and 200 µM limonene. Samples collected in spring 2022. The median is depicted as the middle hinge in the boxplots. Upper and lower hinges represent the first and third quartile. The length of the whispers is determined by the largest and the smallest value in the dataset that are within 1.5 times the interquartile range. (n = 3, *P < .05, **P < .01).

Figure 5.

Log-logistic model fitting to estimate the EC₅₀ and EC₈₀ values of three BNIs in the AS and CS. (A) and (D) log-logistic model for MHPP, (B) and (E) for MHPA, and (C) and (F) for limonene. (A), (B), and (C) correspond to the inhibitors assessed in the AS, and (D), (E), and (F) in the CS. The corresponding EC₅₀, EC₈₀ values (dashed lines), and hill coeficient, with their respective standard errors, as well as the R² are displayed for each BNI in each soil.