Data_Sheet_1_Rice straw increases microbial nitrogen fixation, bacterial and nifH genes abundance with the change of land use types.docx

Soil microorganisms play an important role in soil ecosystems as the main decomposers of carbon and nitrogen. They have an indispensable impact on soil health, and any alterations in the levels of organic carbon and inorganic nitrogen can significantly affect soil chemical properties and microbial community composition. Previous studies have focused on the effects of carbon and nitrogen addition on a single type of soil, but the response of soil microorganisms to varying carbon and nitrogen inputs under different land soil use types have been relatively understudied, leaving a gap in our understanding of the key influencing factors. To address this gap, we conducted a study in the tropical regions of Hainan province, focusing on four distinct land use types: natural forest soil (NS), healthy banana soil (HS), diseased banana garden soil (DS), and paddy soil (PS). Within each of these environments, we implemented five treatments: CK, RS (rice straw), RSN (rice straw and NH4NO3), RR (rice root), and RRN (rice root and NH4NO3). Our aim was to investigate how soil bacteria response to changes in carbon and nitrogen inputs, and to assess their potential for biological nitrogen fixation. The results showed that the addition of rice straw increased the absorption and utilization of nitrate nitrogen by microorganisms. The addition of rice roots (RR) did not increase the absorption capacity of inorganic nitrogen by microorganisms, but increased the content of poorly soluble organic carbon. Most importantly, the addition of rice straw increased microbial respiration and the utilization efficiency of N2 by microorganisms, and the further addition of ammonium nitrate increased microbial respiration intensity. With the change of soil type, the rice straw increases microbial nitrogen fixation, bacterial and nifH genes abundance. Meanwhile, microbial respiration intensity is an important factor influencing the differences in the structure of bacterial communities. The addition of inorganic nitrogen resulted in ammonium nitrogen accumulation, reduced microbial richness and diversity, consequently diminishing the soil microorganisms to resist the environment. Therefore, we believe that with the change of soil types, corresponding soil nutrient retention strategies should be devised and incorporated while reducing the application of ammonium nitrogen, thus ensuring healthy soil development.</p

Data_Sheet_1_Ammonia- and Nitrite-Oxidizing Bacteria are Dominant in Nitrification of Maize Rhizosphere Soil Following Combined Application of Biochar and Chemical Fertilizer.docx

Autotrophic nitrification is regulated by canonical ammonia-oxidizing archaea (AOA) and bacteria (AOB) and nitrite-oxidizing bacteria (NOB). To date, most studies have focused on the role of canonical ammonia oxidizers in nitrification while neglecting the NOB. In order to understand the impacts of combined biochar and chemical fertilizer addition on nitrification and associated nitrifiers in plant rhizosphere soil, we collected rhizosphere soil from a maize field under four different treatments: no fertilization (CK), biochar (B), chemical nitrogen (N) + phosphorus (P) + potassium (K) fertilizers (NPK), and biochar + NPK fertilizers (B + NPK). The potential nitrification rate (PNR), community abundances, and structures of AOA, AOB, complete ammonia-oxidizing bacteria (Comammox Nitrospira clade A), and Nitrobacter- and Nitrospira-like NOB were measured. Biochar and/or NPK additions increased soil pH and nutrient contents in rhizosphere soil. B, NPK, and B + NPK treatments significantly stimulated PNR and abundances of AOB, Comammox, and Nitrobacter- and Nitrospira-like NOB, with the highest values observed in the B + NPK treatment. Pearson correlation and random forest analyses predicted more importance of AOB, Comammox Nitrospira clade A, and Nitrobacter- and Nitrospira-like NOB abundances over AOA on PNR. Biochar and/or NPK additions strongly altered whole nitrifying community structures. Redundancy analysis (RDA) showed that nitrifying community structures were significantly affected by pH and nutrient contents. This research shows that combined application of biochar and NPK fertilizer has a positive effect on improving soil nitrification by affecting communities of AOB and NOB in rhizosphere soil. These new revelations, especially as they related to understudied NOB, can be used to increase efficiency of agricultural land and resource management.</p

Data_Sheet_2_Ammonia- and Nitrite-Oxidizing Bacteria are Dominant in Nitrification of Maize Rhizosphere Soil Following Combined Application of Biochar and Chemical Fertilizer.docx

Autotrophic nitrification is regulated by canonical ammonia-oxidizing archaea (AOA) and bacteria (AOB) and nitrite-oxidizing bacteria (NOB). To date, most studies have focused on the role of canonical ammonia oxidizers in nitrification while neglecting the NOB. In order to understand the impacts of combined biochar and chemical fertilizer addition on nitrification and associated nitrifiers in plant rhizosphere soil, we collected rhizosphere soil from a maize field under four different treatments: no fertilization (CK), biochar (B), chemical nitrogen (N) + phosphorus (P) + potassium (K) fertilizers (NPK), and biochar + NPK fertilizers (B + NPK). The potential nitrification rate (PNR), community abundances, and structures of AOA, AOB, complete ammonia-oxidizing bacteria (Comammox Nitrospira clade A), and Nitrobacter- and Nitrospira-like NOB were measured. Biochar and/or NPK additions increased soil pH and nutrient contents in rhizosphere soil. B, NPK, and B + NPK treatments significantly stimulated PNR and abundances of AOB, Comammox, and Nitrobacter- and Nitrospira-like NOB, with the highest values observed in the B + NPK treatment. Pearson correlation and random forest analyses predicted more importance of AOB, Comammox Nitrospira clade A, and Nitrobacter- and Nitrospira-like NOB abundances over AOA on PNR. Biochar and/or NPK additions strongly altered whole nitrifying community structures. Redundancy analysis (RDA) showed that nitrifying community structures were significantly affected by pH and nutrient contents. This research shows that combined application of biochar and NPK fertilizer has a positive effect on improving soil nitrification by affecting communities of AOB and NOB in rhizosphere soil. These new revelations, especially as they related to understudied NOB, can be used to increase efficiency of agricultural land and resource management.</p

Frequency of the most abundant bacterial genera, indicated in % of all classified sequences, within each treatment of bio-organic fertilizer (BIO), cattle manure compost (CM), Chinese medicine residue compost (CMR), general operation control (GCK) and pig manure compost (PM).

<p>Only the genera frequency higher than 1% was listed in the table. Values are the means followed by standard error of the mean. Different letters indicate statistically significant differences at the 0.05 probability level according to Fisher's least significant difference test (LSD) and the Duncan test.</p

Data_Sheet_1_Impacts of urea and 3,4-dimethylpyrazole phosphate on nitrification, targeted ammonia oxidizers, non-targeted nitrite oxidizers, and bacteria in two contrasting soils.docx

This study explored the effects of combined urea and 3,4-dimethylpyrazole phosphate (DMPP) on several components critical to the soil system: net nitrification rates; communities of targeted ammonia oxidizers [ammonia-oxidizing archaea (AOA) and bacteria (AOB) and complete ammonia-oxidizing bacteria (comammox)]; non-targeted nitrite-oxidizing bacteria (NOB) and bacteria. We conducted the study in two contrasting soils (acidic and neutral) over the course of 28 days. Our results indicated that DMPP had higher inhibitory efficacy in the acidic soil (30.7%) compared to the neutral soil (12.1%). The abundance of AOB and Nitrospira-like NOB were positively associated with nitrate content in acidic soil. In neutral soil, these communities were joined by the abundance of AOA and Nitrobacter-like NOB in being positively associated with nitrate content. By blocking the growth of AOB in acidic soil—and the growth of both AOB and comammox in neutral soil—DMPP supported higher rates of AOA growth. Amplicon sequencing of the 16S rRNA gene revealed that urea and urea + DMPP treatments significantly increased the diversity indices of bacteria, including Chao 1, ACE, Shannon, and Simpson in the acidic soil but did not do so in the neutral soil. However, both urea and urea + DMPP treatments obviously altered the community structure of bacteria in both soils relative to the control treatment. This experiment comprehensively analyzed the effects of urea and nitrification inhibitor on functional guilds involved in the nitrification process and non-targeted bacteria, not just focus on targeted ammonia oxidizers.</p

The relative abundance of the phyla for treatments with bio-organic fertilizer (BIO), cattle manure compost (CM), Chinese medicine residue compost (CMR), general operation control (GCK) and pig manure compost (PM).

<p>Bars represent the standard error of the three replicates and different letters above each phylum indicate significantly difference at 0.05 probability level according to the Duncan test.</p

Rarefaction analysis at different 3% dissimilarity levels for treatments with bio-organic fertilizer (BIO), cattle manure compost (CM), Chinese medicine residue compost (CMR), general operation control (GCK) and pig manure compost (PM).

<p>Rarefaction analysis at different 3% dissimilarity levels for treatments with bio-organic fertilizer (BIO), cattle manure compost (CM), Chinese medicine residue compost (CMR), general operation control (GCK) and pig manure compost (PM).</p

Redundancy analysis (RDA) of the abundant phyla and soil properties for soil samples from treatments with bio-organic fertilizer (BIO), cattle manure compost (CM), Chinese medicine residue compost (CMR), general operation control (GCK) and pig manure compost (PM).

<p>Redundancy analysis (RDA) of the abundant phyla and soil properties for soil samples from treatments with bio-organic fertilizer (BIO), cattle manure compost (CM), Chinese medicine residue compost (CMR), general operation control (GCK) and pig manure compost (PM).</p

Frequency of the most abundant bacterial genera, indicated in % of all classified sequences, within each treatment of bio-organic fertilizer (BIO), cattle manure compost (CM), Chinese medicine residue compost (CMR), general operation control (GCK) and pig manure compost (PM).

<p>Only the genera frequency higher than 1% was listed in the table. Values are the means followed by standard error of the mean. Different letters indicate statistically significant differences at the 0.05 probability level according to Fisher's least significant difference test (LSD) and the Duncan test.</p

Good quality sequences that were used to further analysis after basic quality control for treatments: bio-organic fertilizer (BIO), cattle manure compost (CM), Chinese medicine residue compost (CMR), general operation control (GCK) and pig manure compost (PM).

<p>Good quality sequences that were used to further analysis after basic quality control for treatments: bio-organic fertilizer (BIO), cattle manure compost (CM), Chinese medicine residue compost (CMR), general operation control (GCK) and pig manure compost (PM).</p