The addition of discrimination inhibitors stimulations discrimination potential and N2O emissions were linked to predation among microorganisms in long term nitrogen application and straw returning systems

Abstract

Introduction: Ammonia oxidizing archaea (AOA) and ammonia oxidizing bacteria (AOB) have been proven to be key microorganisms driving the ammonia oxidation process. However, under different fertilization practices, there is a lack of research on the impact of interaction between predators and AOA or AOB on nitrogen cycling at the multi-trophic level.

Methods: In this study, a network-oriented microscopic culture experiment was established based on four different long-term fertilization practices soils. We used the nitrification inhibitors 2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxide-3-oxyl (PTIO) and 3, 4-Dimethylpyrazole phosphate (DMPP) inhibited AOA and AOB, respectively, to explore the impact of interaction between protists and AOA or AOB on nitrogen transformation.

Results: The results showed that long-term nitrogen application promoted the potential nitrification rate (PNR) and nitrous oxide (N₂O) emission, and significantly increased the gene abundance of AOB, but had no obvious effect on AOA gene abundance. DMPP significantly reduced N₂O emission and PNR, while PTIO had no obvious effect on them. Accordingly, in the multi-trophic microbial network, Cercozoa and Proteobacteria were identified as keystone taxa of protists and AOB, respectively, and were significantly positively correlated with N₂O, PNR and nitrate nitrogen. However, Nitrososphaerota archaeon as the keystone species of AOA, had an obvious negative linkage to these indicators. The structural equation model (SEM) showed that AOA and AOB may be competitors to each other. Protists may promote AOB diversity through direct trophic interaction with AOA.

Conclusion: The interaction pattern between protists and ammonia-oxidizing microorganisms significantly affects potential nitrification rate and N₂O emission, which has important implications for soil nitrogen cycle.

Keywords: N2O emission; ammonia-oxidizing bacteria (archaea); keystone taxa; potential nitrification rate; predatory relationship; protists.

FIGURE 1

Dynamic of N₂O emission rates in the four fertilization practices from NN (A), CF (B), SM (C) and SMCF (D) under four treatments, including CK, U, UP and UD during the experimental incubation. Vertical bars indicate standard errors of the means (n = 3). NN, no straw and no nitrogen fertilization; CF, no straw and conventional fertilization; SM, straw mulching and no nitrogen fertilization; SMCF, straw mulching and conventional fertilization. CK, water only; U, urea only; UP, urea + PTIO; UD, urea + DMPP.

FIGURE 2

Dynamic of the AOA (A–D) and AOB (E–H) gene copies in the four fertilization practices from NN (A,E), CF (B,F), SM (C,G), and SMCF (D,H) under four treatments, including CK, U, UP, and UD during the experimental incubation. Vertical bars indicate standard errors of the means, different lowercase letters represent significant differences within groups (n = 3). NN, no straw and no nitrogen fertilization; CF, no straw and conventional fertilization; SM, straw mulching and no nitrogen fertilization; SMCF, straw mulching and conventional fertilization; CK, water only; U, urea only; UP, urea + PTIO; UD, urea + DMPP.

FIGURE 3

The PCoA analysis of protists, AOA and AOB within four fertilization practices (NN, CF, SM, SMCF) under four treatments, including CK, U, UP, and UD. NN, no straw and no nitrogen fertilization; CF, no straw and conventional fertilization; SM, straw mulching and no nitrogen fertilization; SMCF, straw mulching and conventional fertilization; CK, water only; U, urea only; UP, urea + PTIO; UD, urea + DMPP.

FIGURE 4

The co-occurring network of protist, AOA and AOB under four treatments, including CK, U, UP and UD (A). Classification of nodes to identify keystone species within the networks (B). NN, no straw and no nitrogen fertilization; CF, no straw and conventional fertilization; SM, straw mulching and no nitrogen fertilization; SMCF, straw mulching and conventional fertilization; CK, water only; U, urea only; UP, urea + PTIO; UD, urea + DMPP.

FIGURE 5

Pearson correlations between key taxa and physicochemical properties (A) and within key taxa (B). *P < 0.05, **P < 0.01, ***P < 0.001. NN, no straw and no nitrogen fertilization; CF, no straw and conventional fertilization; SM, straw mulching and no nitrogen fertilization; SMCF, straw mulching and conventional fertilization; CK, water only; U, urea only; UP, urea + PTIO; UD, urea + DMPP.

FIGURE 6

Structural equation model (SEM) analysis of the relationships between soil properties, protist, AOA, AOB and N₂O and PNR under CK (A), U (B), UP (C), UD (D) treatments. The arrow width is proportional to the strength of the relationship. Green edge: positive correlation; Red edge: negative correlation. Adjacent numbers in the same direction as the arrow indicate the correlation coefficient. Paths with non-significant coefficients are shown as gray lines. The significance levels are indicated by *P < 0.05, **P < 0.01, ***P < 0.001. NN, no straw and no nitrogen fertilization; CF, no straw and conventional fertilization; SM, straw mulching and no nitrogen fertilization; SMCF, straw mulching and conventional fertilization; CK, water only; U, urea only; UP, urea + PTIO; UD, urea + DMPP.