Response of comammox Nitrospira clades A and B communities to long-term fertilization and rhizosphere effects and their relative contribution to nitrification in a subtropical paddy field of China.

The recently discovered complete ammonia oxidation (comammox Nitrospira) containing clade A and clade B has further complemented our understanding of nitrification process. Nevertheless, understanding the community feature of comammox Nitrospira clades A and B and their relative contribution to nitrification in paddy rhizosphere are still in its infancy. In this study, we assessed the community diversity and structure of comammox Nitrospira clades A and B in paddy rhizosphere and bulk soils under thirty years of different fertilization strategies, i.e., non-fertilization control (CK), chemical fertilizers application (NPK), and NPK plus swine manure (NPKM), respectively. NPKM significantly increased the a-diversity (Chao1 and Shannon indices) of comammox Nitrospira clade A and altered the community structure (P < 0.05) but had little effect on clade B. A two-way analysis of variance (ANOVA) showed that the effect of long-term fertilization on soil comammox Nitrospira community and nitrification potential rate (PNR) was much greater than that of rhizosphere. Compared with NPK, soil PNR was greatly increased by 51.0% under the NPKM treatment in the rhizosphere (P < 0.05). Phylogenetic analysis showed that NPKM improved the relative abundances of sub-clade A.2.1 and sub-clade A.3.2 of the comammox clade A community, with an average increase of 212.2 and 210.4% in both rhizosphere and bulk soils relative to the NPK treatment. Soil organic matter, NH4+-N, and pH were significant soil drivers of comammox Nitrospira clades A and B community. Furthermore, linear regression and structural equation modeling clearly showed that comammox Nitrospira clade A a-diversity were significantly associated with soil PNR (P < 0.05). Our results suggest (i) that comammox Nitrospira clade A are sensitive to the organic fertilization; and (ii) that comammox Nitrospira clade A contribute more to nitrification than clade B under the long-term organic fertilized paddy soil.

Long-term effects of nitrogen and phosphorus fertilization on profile distribution and characteristics of dissolved organic matter in fluvo-aquic soil.

Dissolved organic matter (DOM) drives numerous biogeochemical processes (e.g. carbon cycling) in agro-ecosystems and is sensitive to fertilization management. Nevertheless, changes in the quantity and quality of DOM in the vertical soil profile following long-term continuous nitrogen (N) and phosphorus (P) inputs remain unclear. In this study, the contents and optical characteristics of DOM along a 2-m soil profile were investigated using a 40-year wheat/maize rotation combined with experiments using different N and P fertilization rates in the North China Plain. The results revealed that the dissolved organic carbon (DOC) content decreased with an increase in soil depths. Compared with that in the control (no fertilization), 40-year N, P, and N + P additions increased the soil DOC content by 26%-69%, except for 270-kg N, and 67.5-kg P treatments. N + P application resulted in higher DOC contents than N-alone and P-alone applications. N, P, and N + P inputs increased or did not affect the aromaticity and hydrophobicity of DOM at 0-40 cm but reduced them from 40 to 200 cm. Compared with that in the control, N, P, and N + P inputs enhanced the content of humic acid-like substances (C1+C2+C3+C4) and decreased the content of protein-like substance (C5). C1 was the dominant component among the five DOM, representing the microbial humic component. Optical indices also indicated that soil DOM primarily originated from microbial sources. Nutrient addition accelerated transformation between complex C1 and simple C5 via promoting microbial activities. These results imply that N and P fertilizers increased the DOM content and altered its composition, thereby potentially affecting the stability of soil organic matter in the agroe-cosystems.

Contrasting effects of iron oxides on soil organic carbon accumulation in paddy and upland fields under long-term fertilization.

Active iron oxides, especially poorly crystalline forms, benefit soil organic carbon (SOC) accumulation via directly bounding and indirectly promoting aggregation. However, it remains unclear on the impacts of active iron oxides on SOC accumulation in paddy and upland soils under long-term fertilization regimes. Here, we attempted to clarify the underlying mechanisms of amorphous (FeO) and organically complexed (FeP) iron oxides mediating SOC accumulation in paddy and upland soils based on two long-term fertilization experiments (both including no fertilization [CK]; chemical nitrogen, phosphorus and potassium [NPK] and NPK plus manure [NPKM] treatments). Results showed that compared to upland soil, Fe-bound organic carbon (Fe-bound OC) content in paddy soil, occupying 21-30% of SOC, was 77% higher on average, due to larger amounts of FeO (+31%) and FeP (+224%). The FeO and FeP were positively related to mean weight diameter (MWD) of soil aggregates across paddy and upland soils. Compared to NPK treatment, NPKM treatment strongly increased FeO (+41%), FeP (+60%) and associated Fe-bound OC (+19%) in paddy soil, and increased FeO (+17%) and FeP (+25%) while decreasing Fe-bound OC (-9%) in upland soil. These combined findings indicated the importance of poorly crystalline iron oxides facilitating Fe-bound OC formation and its contribution to SOC accumulation in paddy soil rather than upland soil. Moreover, long-term manure amendment could enhance SOC accumulation by increasing Fe-bound OC and aggregation stability in paddy soil and enhancing physical protection in upland soil, largely attributed to increased poorly crystalline iron oxides. Overall, these results highlight the potential mechanisms through which active iron oxides regulate SOC accumulation and guide fertilization management in paddy and upland soils.

Phosphorus Fertilization Boosts Mineral-Associated Soil Organic Carbon Formation Associated with Phagotrophic Protists.

Long-term fertilization affects soil organic C accumulation. A growing body of research has revealed critical roles of bacteria in soil organic C accumulation, particularly through mineral-associated organic C (MAOC) formation. Protists are essential components of soil microbiome, but the relationships between MAOC formation and protists under long-term fertilization remain unclear. Here, we used cropland soil from a long-term fertilization field trial and conducted two microcosm experiments with 13C-glucose addition to investigate the effects of N and P fertilizations on MAOC formation and the relationships with protists. The results showed that long-term fertilization (especially P fertilization) significantly (P < 0.05) increased 13C-MAOC content. Compared with P-deficient treatment, P replenishment enriched the number of protists (mainly Amoebozoa and Cercozoa) and bacteria (mainly Acidobacteriota, Bacteroidota, and Gammaproteobacteria), and significantly (P < 0.001) promoted the abundances of bacterial functional genes controlling C, N, P, and S metabolisms. The community composition of phagotrophic protists prominently (P < 0.001) correlated with the bacterial community composition, bacterial functional gene abundance, and 13C-MAOC content. Co-occurrence networks of phagotrophic protists and bacteria were more connected in soil with the N inoculum added than in soil with the NP inoculum added. P replenishment strengthened bacterial 13C assimilation (i.e., 13C-phospholipid fatty acid content), which negatively (P < 0.05) correlated with the number and relative abundance of phagotrophic Cercozoa. Together, these results suggested that P fertilization boosts MAOC formation associated with phagotrophic protists. Our study paves the way for future research to harness the potential of protists to promote belowground C accrual in agroecosystems.

Fertilization Weakens the Ecological Succession of Dissolved Organic Matter in Paddy Rice Rhizosphere Soil at the Molecular Level.

Dissolved organic matter (DOM) is involved in numerous biogeochemical processes, and understanding the ecological succession of DOM is crucial for predicting its response to farming (e.g., fertilization) practices. Although plentiful studies have examined how fertilization practice affects the content of soil DOM, it remains unknown how long-term fertilization drives the succession of soil DOM over temporal scales. Here, we investigated the succession of DOM in paddy rice rhizosphere soils subjected to different long-term fertilization treatments (CK: no fertilization; NPK: inorganic fertilization; OM: organic fertilization) along with plant growth. Our results demonstrated that long-term fertilization significantly promoted the molecular chemodiversity of DOM, but it weakened the correlation between DOM composition and plant development. Time-decay analysis indicated that the DOM composition had a shorter halving time under CK treatment (94.7 days), compared to NPK (337.4 days) and OM (223.8 days) treatments, reflecting a lower molecular turnover rate of DOM under fertilization. Moreover, plant development significantly affected the assembly process of DOM only under CK, not under NPK and OM treatments. Taken together, our results demonstrated that long-term fertilization, especially inorganic fertilization, greatly weakens the ecological succession of DOM in the plant rhizosphere, which has a profound implication for understanding the complex plant-DOM interactions.

Microbial Multitrophic Communities Drive the Variation of Antibiotic Resistome in the Gut of Soil Woodlice (Crustacea: Isopoda).

Multitrophic communities inhabit in soil faunal gut, including bacteria, fungi, and protists, which have been considered a hidden reservoir for antibiotic resistance genes (ARGs). However, there is a dearth of research focusing on the relationships between ARGs and multitrophic communities in the gut of soil faunas. Here, we studied the contribution of multitrophic communities to variations of ARGs in the soil woodlouse gut. The results revealed diverse and abundant ARGs in the woodlouse gut. Network analysis further exhibited strong connections between key ecological module members and ARGs, suggesting that multitrophic communities in the keystone ecological cluster may play a pivotal role in the variation of ARGs in the woodlouse gut. Moreover, long-term application of sewage sludge significantly altered the woodlice gut resistome and interkingdom communities. The variation portioning analysis indicated that the fungal community has a greater contribution to variations of ARGs than bacterial and protistan communities in the woodlice gut after long-term application of sewage sludge. Together, our results showed that changes in gut microbiota associated with agricultural practices (e.g., sewage sludge application) can largely alter the gut interkingdom network in ecologically relevant soil animals, with implications for antibiotic resistance, which advances our understanding of the microecological drivers of ARGs in terrestrial ecosystem.

Enhanced phosphorus mobility in a calcareous soil with organic amendments additions: Insights from a long term study with equal phosphorus input.

The agricultural practice of replacing chemical fertilizers with organic amendments (manure and/or straw) may have consequences for phosphorus (P) loss to the environment. Such a knowledge gap was examined using a ten-year field trial in calcareous soil containing four treatments with the equal annual P input but varied organic amendment combinations as follows: mineral fertilizer only as control (MF), mineral fertilizer coupled with manure (MM), mineral fertilizer coupled with manure and straw (MMS) and mineral fertilizer coupled with straw (MS). The soil P distribution, P fractions and speciation, Fe(III) reduction and P sorption kinetics were investigated using the chemical extraction, K edge X-ray absorption near-edge structure and Langmuir equations. The electronic shuttle capacity of soils and speciation of soil dissolved organic matter (DOM) were also evaluated using electrochemical methods, three-dimensional excitation-emission matrix fluorescence spectroscopy and Fourier transform infrared spectra methods. Results showed that soil Olsen-P and total P increased at depths of 20-40 cm in MM, MMS and MS treatments, suggesting that manure and/or straw addition significantly mobilized P in the soil profile. Manure and/or straw addition also decreased soil maximum P sorption capacity (Smax) and increased the desorption rate at depths of 0-20 cm in soil across treatments. At a depth of 0-20 cm in soil of the MS treatment, the enhanced Fe(Ⅲ) reduction coupled with a decrease of Fe-bound P supports that Fe reduction dominates the mobilization of P. The transformation of Ca bound-P to Al/Fe bound-P in a depth of 0-20 cm in soil of the MM treatment may be due to the high proportion of humic-like substances in the DOM at a depth of 0-20 cm in soil of the MM treatment, which may have caused a slight/microsite acidification. These results can help to develop optimized fertilization practices to effectively mitigate P loss from calcareous soils with manure and/or straw addition.

Differential Responses of Arbuscular Mycorrhizal Fungal Communities to Long-Term Fertilization in the Wheat Rhizosphere and Root Endosphere.

Arbuscular mycorrhizal fungi (AMF) provide essential nutrients to crops and are critically impacted by fertilization in agricultural ecosystems. Understanding shifts in AMF communities in and around crop roots under different fertilization regimes can provide important lessons for improving agricultural production and sustainability. Here, we compared the responses of AMF communities in the rhizosphere (RS) and root endosphere (ES) of wheat (Triticum aestivum) to different fertilization treatments, nonfertilization (control), mineral fertilization only (NPK), mineral fertilization plus wheat straw (NPKS), and mineral fertilization plus cow manure (NPKM). We employed high-throughput amplicon sequencing and investigated the diversity, community composition, and network structure of AMF communities to assess their responses to fertilization. Our results elucidated that AMF communities in the RS and ES respond differently to fertilization schemes. Long-term NPK application decreased the RS AMF alpha diversity significantly, whereas additional organic amendments (straw or manure) had no effect. In contrast, NPK fertilization increased the ES AMF alpha diversity significantly, while additional organic amendments decreased it significantly. The effect of different fertilization schemes on AMF network complexity in the RS and ES were similar to their effects on alpha diversity. Changes to AMF communities in the RS and ES correlated mainly with the pH and phosphorus level of the rhizosphere soil under long-term inorganic and organic fertilization regimes. We suggest that the AMF community in the roots should be given more consideration when studying the effects of fertilization regimes on AMF in agroecosystems. IMPORTANCE Arbuscular mycorrhizal fungi are an integral component of rhizospheres, bridging the soil and plant systems and are highly sensitive to fertilization. However, surprisingly little is known about how the response differs between the roots and the surrounding soil. Decreasing arbuscular mycorrhizal fungal diversity under fertilization has been reported, implying a potential reduction in the mutualism between plants and arbuscular mycorrhizal fungi. However, we found opposing responses to long-term fertilization managements of arbuscular mycorrhizal fungi in the wheat roots and rhizosphere soil. These results suggested that changes in the arbuscular mycorrhizal fungal community in soils do not reflect those in the roots, highlighting that the root arbuscular mycorrhizal fungal community is pertinent to understand arbuscular mycorrhizal fungi and their crop hosts' responses to anthropogenic influences.

Molybdenum Bioavailability and Asymbiotic Nitrogen Fixation in Soils are Raised by Iron (Oxyhydr)oxide-Mediated Free Radical Production.

Nitrogen (N) fixation in soils is closely linked to microbially mediated molybdenum (Mo) cycling. Therefore, elucidating the mechanisms and factors that affect Mo bioavailability is crucial for understanding N fixation. Here, we demonstrate that long-term (26 years) manure fertilization increased microbial diversity and content of short-range ordered iron (oxyhydr)oxides that raised Mo bioavailability (by 2.8 times) and storage (by ∼30%) and increased the abundance of nifH genes (by ∼14%) and nitrogenase activity (by ∼60%). Nanosized iron (oxyhydr)oxides (ferrihydrite, goethite, and hematite nanoparticles) play a dual role in soil Mo cycling: (i) in concert with microorganisms, they raise Mo bioavailability by catalyzing hydroxyl radical (HO•) production via the Fenton reactions and (ii) they increase Mo retention by association with the nanosized iron (oxyhydr)oxides. In summary, long-term manure fertilization raised the stock and bioavailability of Mo (and probably also of other micronutrients) by increasing iron (oxyhydr)oxide reactivity and intensified asymbiotic N fixation through an increased abundance of nifH genes and nitrogenase activity. This work provides a strategy for increasing biological N fixation in agricultural ecosystems.

Effects of nearly four decades of long-term fertilization on the availability, fraction and environmental risk of cadmium and arsenic in red soils.

Fertilizers are important for agricultural production because they can effectively promote crop productivity. However, long-term fertilization can cause heavy metal accumulation in soils and crops. This study utilized sequential extraction, the diffusive gradient in the thin films (DGT) technique and risk assessment models to estimate the effects of the longest long-term fertilization (38 years) in China on cadmium (Cd) and arsenic (As) accumulation in soils. The treatments included no fertilization (CK); inorganic nitrogen, phosphorus, and potassium fertilization (NPK); manure fertilization (M); and NPK plus M cofertilization (NPKM). The results indicated that the soils treated with NPKM, M and NPK had significantly increased total and available concentrations of Cd and As after 38 years of long-term fertilization. Cd mainly originates from cattle manure, while As originates from phosphate fertilizer. Sequential extraction results indicated that the application of manure increased the acid/exchangeable fraction (F1) and organic matter-bound fraction (F3) of Cd and As. The risk assessment results showed that the environmental risks of both Cd and As increased during long-term fertilization, and Cd contamination in the soil was at a moderate-high level, while As remained at a relatively low level. According to the calculations of the maximum numbers of years of soil productivity and rice production, Cd was labile and accumulated in the soils, and As was more labile than Cd in terms of accumulating in rice, indicating that the true risk from As in rice is higher than that from Cd. Controlling the heavy metals in fertilizers, mitigating effective amendments, and identifying plant types that accumulate low amounts of contaminants may be good choices for cleaner crop production.

Soil aggregation and soil aggregate stability regulate organic carbon and nitrogen storage in a red soil of southern China.

Soil aggregation plays a critical role in the maintenance of soil structure, as well as in its productivity. Fertilization influences soil aggregation, especially by regulating soil organic carbon (SOC) and total nitrogen (TN) contents in aggregate fractions. The present study evaluated the influence of three contrasting fertilizer regimes (unfertilized control -CK-, mineral fertilization -NPK- and manure combined with NPK -NPKM) on soil aggregate stability, aggregate-associated organic carbon and total nitrogen sequestration and mineralization of SOC. Soil samples from (20 cm) depth were collected from a long-term fertilization experiment and analysed for size distribution ranging (>250 µm, 250-53 µm and <53 µm sizes), SOC and TN contents, as well as for mineralization of bulk and aggregate associated-SOC. Both NPK and NPKM fertilizations significantly enhanced SOC and TN contents in bulk soil and its constituent aggregates of >250 µm, 250-53 µm and <53 µm sizes, as compared to CK. Long-term NPK and NPKM increased SOC and TN stock in bulk soil by 45 and 98%, and by 70 and 144%, respectively, as compared to CK. Similarly, higher values of SOC and TN stock in all aggregate fractions was observed with the application of NPKM. Application of NPK and NPKM for 26 years significantly increased aggregate stability, which was positively correlated with total SOC contents in terms of mean weight diameter (MWD) (Adj. R2 = 0.689, p < 0.03) and geometric mean diameter (GMD) (Adj. R2 = 0.471, p < 0.24). Moreover, higher scores regarding cumulative mineralization for bulk soil and aggregate associated OC were observed with the application of NPK and NPKM. Irrespective of treatments, higher cumulative C-mineralization was observed for macro-aggregates (>250 µm size) followed by 250-53 µm and <53 µm size aggregates. Interestingly, a highly positive correlation was observed between aggregate stability and the cumulative amount of mineralization for bulk soil and aggregate fractions, with R2 ranging from 0.84 to 0.99. This study evidenced that long-term fertilization of NPK and NPKM can improve soil aggregation, stability and associated OC and TN stock in aggregates, as well as aggregate-associated OC mineralization, which was further governed by aggregate size.

Utilization of carbon sources in the rice rhizosphere and nonrhizosphere soils with different long-term fertilization management.

Carbon (C) plays an important role in the interaction between plant and rhizosphere microbial communities, but there is still limited information about how C source utilization soil microbial structure responds to soil fertility changes under the double-cropping rice (Oryza sativa L.) system in Southern China paddy fields. Therefore, the effects of long-term (33 years) fertilizer regimes on the characteristics of C utilization in both rhizosphere and nonrhizosphere soils under double-cropping rice fields in Southern China were investigated by using the metagenome sequencing technology. The experiment began in 1986, and included five fertilizer treatments: without fertilizer input (CK), chemical fertilizer alone (MF), rice straw residue and chemical fertilizer (RF), 30% organic matter, and 70% chemical fertilizer (LOM), and 60% organic matter and 40% chemical fertilizer (HOM). The results showed that the relative abundance of Gemmatimonadetes and Planctomycetia in both the rhizosphere and nonrhizosphere soils was increased by application of rice straw residue and organic manure, whereas the relative abundance of Gammaproteobacteria and Nitrospira was promoted by application of inorganic fertilizers. The largest group of clusters of orthologous groups of proteins categories was "amino acid transport and metabolism" with 16.46% unigenes, followed by "general function prediction only" (12.23%). Regarding the gene ontology categories, biological process were the largest category (174 949, 46.40%), followed by cellular component (126 766, 33.62%), and molecular function (110 353, 29.26%). The principal coordinate analysis indicated that different parts of the root zone were the most important factors affecting the variation of C source utilization bacteria community, and the different fertilizer treatments were the second important factor affecting the variation of C source utilization bacteria community. As a result, the application of fertilization practices had significant effects on the abundance and community composition of C source utilization microbes in paddy soils. The results showed that the combined application of rice straw residue or organic manure with chemical fertilizer practices significantly increases the C source utilization of soil microorganisms in double-cropping rice fields.

Nitrosospira Cluster 8a Plays a Predominant Role in the Nitrification Process of a Subtropical Ultisol under Long-Term Inorganic and Organic Fertilization.

Long-term effects of inorganic and organic fertilization on nitrification activity (NA) and the abundances and community structures of ammonia-oxidizing bacteria (AOB) and ammonia-oxidizing archaea (AOA) were investigated in an acidic Ultisol. Seven treatments applied annually for 27 years comprised no fertilization (control), inorganic NPK fertilizer (N), inorganic NPK fertilizer plus lime (CaCO3) (NL), inorganic NPK fertilizer plus peanut straw (NPS), inorganic NPK fertilizer plus rice straw (NRS), inorganic NPK fertilizer plus radish (NR), and inorganic NPK fertilizer plus pig manure (NPM). In nonfertilized soil, the abundance of AOA was 1 order of magnitude higher than that of AOB. Fertilization reduced the abundance of AOA but increased that of AOB, especially in the NL treatment. The AOA communities in the control and the N treatments were dominated by the Nitrososphaera and B1 clades but shifted to clade A in the NL and NPM treatments. Nitrosospira cluster 8a was found to be the most dominant AOB in all treatments. NA was primarily regulated by soil properties, especially soil pH, and the interaction with AOB abundance explained up to 73% of the variance in NA. When NL soils with neutral pH were excluded from the analysis, AOB abundance, especially the relative abundance of Nitrosospira cluster 8a, was positively associated with NA. In contrast, there was no association between AOA abundance and NA. Overall, our data suggest that Nitrosospira cluster 8a of AOB played an important role in the nitrification process in acidic soil following long-term inorganic and organic fertilization.IMPORTANCE The nitrification process is an important step in the nitrogen (N) cycle, affecting N availability and N losses to the wider environment. Ammonia oxidation, which is the first and rate-limiting step of nitrification, was widely accepted to be mainly regulated by AOA in acidic soils. However, in this study, nitrification activity was correlated with the abundance of AOB rather than that of AOA in acidic Ultisols. Nitrosospira cluster 8a, a phylotype of AOB which preferred warm temperatures, and low soil pH played a predominant role in the nitrification process in the test Ultisols. Our results also showed that long-term application of lime or pig manure rather than plant residues altered the community structure of AOA and AOB. Taken together, our findings contribute new knowledge to the understanding of the nitrification process and ammonia oxidizers in subtropical acidic Ultisol under long-term inorganic and organic fertilization.

Microbial composition and diversity are associated with plant performance: a case study on long-term fertilization effect on wheat growth in an Ultisol.

The association between microbial communities and plant growth in long-term fertilization system has not been fully studied. In the present study, impacts of long-term fertilization have been determined on the size and activity of soil microbial communities and wheat performance in a red soil (Ultisol) collected from Qiyang Experimental Station, China. For this, different microbial communities originating from long-term fertilized pig manure (M), mineral fertilizer (NPK), pig manure plus mineral fertilizer (MNPK), and no fertilizer (CK) were used as inocula for the Ultisol tested. Changes in total bacterial and fungal community composition and structures using Ion Torrent sequencing were determined. The results show that the biomass of wheat was significantly higher in both sterilized soil inoculated with NPK (SNPK) and sterilized soil inoculated with MNPK (SMNPK) treatments than in other treatments (P < 0.05). The activities of ß-1,4-N-acetylglucosaminidase (NAG) and cellobiohydrolase (CBH) were significantly correlated with wheat biomass. Among the microbial communities, the largest Ascomycota phylum in soils was negatively correlated with ß-1,4-glucosidase (ßG) (P < 0.05). The phylum Basidiomycota was negatively correlated with plant biomass (PB) and tillers per plant (TI) (P < 0.05). Nonmetric multidimensional scaling analysis shows that fungal community was strongly correlated with long-term fertilization strategy, while the bacterial community was strongly correlated with ß-1,4-N-acetylglucosaminidase activity. According to the Mantel test, the growth of wheat was affected by fungal community. Taken together, microbial composition and diversity in soils could be a good player in predicting soil fertility and consequently plant growth.

Long-term manure amendments and chemical fertilizers enhanced soil organic carbon sequestration in a wheat (Triticum aestivum L.)-maize (Zea mays L.) rotation system.

BACKGROUND: The carbon sequestration potential is affected by cropping system and management practices, but soil organic carbon (SOC) sequestration potential under fertilizations remains unclear in north China. This study examined SOC change, total C input to soil and, via integration of these estimates over years, carbon sequestration efficiency (CSE, the ratio of SOC change over C input) under no fertilization (control), chemical nitrogen fertilizer alone (N) or combined with phosphorus and potassium fertilizers (NP, NK, PK and NPK), or chemical fertilizers combined with low or high (1.5×) manure input (NPKM and 1.5NPKM). RESULTS: Results showed that, as compared with the initial condition, SOC content increased by 0.03, 0.06, 0.05, 0.09, 0.16, 0.26, 0.47 and 0.68 Mg C ha-1 year-1 under control, N, NK, PK, NP, NPK, NPKM and 1.5NPKM treatments respectively. Correspondingly, the C inputs of wheat and maize were 1.24, 1.34, 1.55, 1.33, 2.72, 2.96, 2.97 and 3.15 Mg ha-1 year-1 respectively. The long-term fertilization-induced CSE showed that about 11% of the gross C input was transformed into SOC pool. CONCLUSION: Overall, this study demonstrated that decade-long manure input combined with chemical fertilizers can maintain high crop yield and lead to SOC sequestration in north China. © 2016 Society of Chemical Industry.

Long-term balanced fertilization increases the soil microbial functional diversity in a phosphorus-limited paddy soil.

The influence of long-term chemical fertilization on soil microbial communities has been one of the frontier topics of agricultural and environmental sciences and is critical for linking soil microbial flora with soil functions. In this study, 16S rRNA gene pyrosequencing and a functional gene array, geochip 4.0, were used to investigate the shifts in microbial composition and functional gene structure in paddy soils with different fertilization treatments over a 22-year period. These included a control without fertilizers; chemical nitrogen fertilizer (N); N and phosphate (NP); N and potassium (NK); and N, P and K (NPK). Based on 16S rRNA gene data, both species evenness and key genera were affected by P fertilization. Functional gene array-based analysis revealed that long-term fertilization significantly changed the overall microbial functional structures. Chemical fertilization significantly increased the diversity and abundance of most genes involved in C, N, P and S cycling, especially for the treatments NK and NPK. Significant correlations were found among functional gene structure and abundance, related soil enzymatic activities and rice yield, suggesting that a fertilizer-induced shift in the microbial community may accelerate the nutrient turnover in soil, which in turn influenced rice growth. The effect of N fertilization on soil microbial functional genes was mitigated by the addition of P fertilizer in this P-limited paddy soil, suggesting that balanced chemical fertilization is beneficial to the soil microbial community and its functions.

Influence of long-term fertilization on soil microbial biomass, dehydrogenase activity, and bacterial and fungal community structure in a brown soil of northeast China.

In this study, the effect of mineral fertilizer and organic manure were evaluated on soil microbial biomass, dehydrogenase activity, bacterial and fungal community structure in a long-term (33 years) field experiment. Except for the mineral nitrogen fertilizer (N) treatment, long-term fertilization greatly increased soil microbial biomass carbon (SMBC) and dehydrogenase activity. Organic manure had a significantly greater impact on SMBC and dehydrogenase activity, compared with mineral fertilizers. Bacterial and fungal community structure was analyzed by polymerase chain reaction (PCR)-denaturing gradient gel electrophoresis (DGGE). Long-term fertilization increased bacterial and fungal ribotype diversity. Total soil nitrogen (TN) and phosphorus (TP), soil organic carbon (SOC) and available phosphorus (AP) had a similar level of influence on bacterial ribotypes while TN, SOC and AP had a larger influence than alkali-hydrolyzable nitrogen (AHN) on fungal ribotypes. Our results suggested that long-term P-deficiency fertilization can significantly decrease soil microbial biomass, dehydrogenase activity and bacterial diversity. N-fertilizer and SOC have an important influence on bacterial and fungal communities.

[Effects of Long-term Fertilizations on the Organic Carbon Components of Soil Macroaggregates and the Yield of Wheat in Wheat Fields on the Loess Plateau].

Soil macro-aggregates are the main location for soil organic carbon （SOC） sequestration, which is of great significance to improve soil fertility. This study aimed to understand the mechanisms of the organic carbon （OC） sequestration in macroaggregates and improve crop yield in wheat fields on the loess plateau. With the aggregate-density fractionation method, an eight-year experiment was conducted to investigate the following three factors： ① the effects of long-term fertilization on OC fractions within macroaggregates； ② the variation characteristics of OC fractions within macroaggregates, including coarse particulate organic carbon （cPOC）, fine particulate organic carbon （fPOC）, intra-microaggregate particulate organic carbon （iPOC）, free silt and clay particulate carbon （s+c_f）, and intra-microaggregate silt and clay particulate carbon （s+c_m）； ③ and the relationships between them and SOC input and yield formation. The treatments included no fertilization （CK）, farmer pattern （NP）, optimized fertilizers pattern （NPK）, optimized fertilizers + organic fertilizers pattern （NPKM）, and optimized fertilizers + biological organic fertilizers pattern （NPKB）. The results showed that the application of organic and chemical fertilizer （NPKM and NPKB） improved significantly the SOC content in macroaggregates compared with that in the single fertilizer treatment （NP and NPK）, which had a greater increase in SOC content in macroaggregates than that of the soil. All fertilization treatments had a tendency to increase the content of fractions iPOC, fPOC, and iPOC in macroaggregates, but silt and clay carbon （s+c_f and s+c_m） contents were decreased. The application of manure combined with chemicals markedly increased the allocations of fractions cPOC, fPOC, and iPOC reserves, but it greatly decreased （s+c_f） reserves allocation. However, the application of chemical fertilizers only significantly increased the proportion of cPOC reserves in macroaggregates. Correlation analysis showed that there were significant positive correlations among wheat grain yield and OC fractions （cPOC and fPOC） contents, SOC content, the OC content of >0.25 mm macroaggregates, and SOC input, and the correlation coefficient was 0.645-0.883. In conclusion, long-term fertilization, especially combined with organic fertilizer, could promote the free silt and clay carbon fraction （s+c_f） to transfer into other forms of OC components through the increase in soil carbon input in the wheat field of the loess plateau. Furthermore, the OC content of macroaggregates was increased overall, providing a good soil environment for crop yield.

Ensemble learning algorithms to elucidate the core microbiome's impact on carbon content and degradation properties at the soil aggregate level.

Soil aggregates are crucial for soil organic carbon (OC) accumulation. This study, utilizing a 32-year fertilization experiment, investigates whether the core microbiome can elucidate variations in carbon content and decomposition across different aggregate sizes more effectively than broader bacterial and fungal community analyses. Employing ensemble learning algorithms that integrate machine learning with network inference, we found that the core microbiome accounts for an average increase of 26 % and 20 % in the explained variance of PCoA and Adonis analyses, respectively, in response to fertilization. Compared to the control, inorganic and organic fertilizers decreased the decomposition index (DDI) by 31 % and 38 %, respectively. The fungal core microbiome predominantly influenced OC content and DDI in larger macroaggregates (>2000 µm), explaining over 35 % of the variance, while the bacterial core microbiome had a lesser impact, explaining <30 %. Conversely, in smaller aggregates (<2000 µm), the bacterial core microbiome significantly influenced DDI (R2 > 0.2), and the fungal core microbiome more strongly affected OC content (R2 > 0.3). Mantel tests showed that pH is the most significant environmental factor affecting core microbiome composition across all aggregate sizes (Mantel's r > 0.8, P < 0.01). Linear correlation analysis further confirmed that the core microbiome's community structure could accurately predict OC content and DDI in aggregates (R2 > 0.8, P < 0.05). Overall, our findings suggested that the core microbiome provides deeper insights into the variability of aggregate organic carbon content and decomposition, with the bacterial core microbiome playing a particularly pivotal role within the soil aggregates.

[Effects of 40 Years of Fertilizer Application on the Characteristics of Soil Enzymatic Stoichiometry in Black Soil].

Microorganisms produce extracellular enzymes to meet elemental requirements and cope with stoichiometric imbalances of resources. To gain insights into the cycling of C, N, and P, the activities of the C∶N∶P acquisition enzymes have been extensively investigated. To detect the effects of long-term fertilization practices on soil nutrient balance and characteristics of soil enzymatic stoichiometry in black soil, four different fertilization treatments were selected： no fertilization （CK）, nitrogen fertilizer （N）, phosphorus fertilizer （P）, and combination of nitrogen and phosphorus fertilizers （NP）. Soil samples were collected in both April 2021 and April 2022 to determine soil enzyme activities and their stoichiometric characteristics. The results showed that soil acid phosphatase and ß-D-glucosidase activities were significantly higher in the N and NP treatments than in CK by 68%-158% and 26%-222%, respectively. Soil ß-N-acetylaminoglucosidase activities were significantly higher in the P and NP treatments, with the highest around 75.48 nmol·ï¼ˆg·h）-1 and 106.81 nmol·ï¼ˆg·h）-1, respectively. Two-way ANOVA analysis showed that N and P inputs had a great impact on soil enzyme activities. Redundancy analysis showed that the main factors controlling enzyme activities were soil pH, microbial biomass phosphorus, and soil available P content. It was found that N inputs significantly increased enzyme vector length, which was ranged from 1.32 to 1.52, and the enzyme vector angles were all larger than 45°, suggesting C and P co-limited in the black soils. These findings suggest that 40 years of fertilization have had a great impact on soil enzymes and the related resource use strategy, which provides great implications for assessing soil nutrients balance and soil sustainability.