Soil organic matter (SOM) is crucial for ecosystem carbon cycling, soil fertility, and environmental quality. As the main component of SOM, humic substances (HS) are considered a unique category of nonuniformly assembled substances. It is widely accepted that HS are originated from small molecules produced during the decomposition of plant and animal residues or from residual macromolecules. These molecules can be recombined or condensed via enzymatic and mineral catalysis into quasi-macromolecular compounds or compound groups with high condensation (elevated C/H molar ratio) and relatively large molecular weight and are further stabilized by mineral association. Although HS can be regarded as an extension or a narrow definition of SOM, their properties are inherently more complex, and their chemical composition, structure, and formation processes remain controversial. Here, we examine the formation theories, compositional structure, stabilization mechanisms, and functional roles of SOM and HS. We propose that HS indeed differ from non-HS, with HS consisting of both residual and synthetic quasi-macromolecular substance components, which collectively form unique compounds or compound groups with independent characteristics. Consequently, HS remain fundamental to soil science and continue to be widely utilized through various HS-based technologies and products in agriculture and environmental fields.

Biobased biodegradable microplastics (Bio-MPs) can alter both the quantity and molecular composition of dissolved organic matter (DOM) in soil, which profoundly shapes the stability of soil organic matter (SOM). However, the microbial mechanisms underlying the Bio-MP-induced DOM turnover, particularly the role of fungi, remain largely unclear. Here, we tracked DOM molecular dynamics using Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) and characterized microbial communities using 16S rRNA gene and ITS amplicon sequencing during a 30-day soil incubation amended with polylactic acid (PLA-MPs, low biodegradability) and polyhydroxyalkanoate (PHA-MPs, high biodegradability). Our results showed that PLA-MPs exerted minimal impacts on DOM dynamics, whereas PHA-MPs rapidly increased DOM content and CO2 emission and shifted the DOM molecular composition from recalcitrant compounds (e.g., lignins and tannins) toward labile compounds (e.g., lipids and proteins/amino sugars). These alterations were primarily driven by fungal depolymerization of PHA-MPs and SOM to generate labile DOM, followed by bacterial assimilation, indicating a fungal-initiated metabolic cascade that governs soil DOM turnover under PHA-MP exposure. The increase in labile DOM was mainly associated with enrichment of fast-growing fungi (e.g., Neocosmospora). Overall, this study elucidates the pivotal role of fungi in mediating Bio-MP-induced DOM turnover and shaping SOM stability.

Background and Aims: Decomposed organic amendments can alter soil nutrient availability and root metabolism. However, whether different amendment types differentially influence these processes to shape rhizosphere bacterial community assembly remains unclear. Methods A pot experiment was conducted using maize ( Zea mays L.) grown in unamended soil (CK), soil amended with decomposed maize straw (S), and soil amended with co-decomposed maize straw and pig manure (SM). Root metabolic profiles and rhizosphere bacterial communities were characterized using untargeted liquid chromatography–mass spectrometry (LC–MS) and 16S ribosomal RNA gene sequencing, respectively. Results Compared with S, SM increased total phosphorus (TP) and available phosphorus (AP) by 18.6% and 37.0%, respectively ( P < 0.05), whereas available potassium (AK) was 20.0% higher under S than under SM. Treatment effects explained 38.26% of root metabolome variation. The composition of the rhizosphere bacterial community differed significantly among the CK, S, and SM ( R ² = 0.346, P = 0.0044). S treatment enriched Chryseolinea , Ruminiclostridium , and Lysinibacillus , whereas SM treatment enriched Ureibacillus , Ohtaekwangia , UCG-012 , and Chryseotalea . Partial least squares path modeling (PLS-PM) indicated that decomposed organic amendments were positively associated with soil nutrient status ( β = 0.95), which was subsequently positively associated with root metabolic reprogramming ( β = 0.89) and bacterial community reorganization ( β = 1.44). In contrast, root metabolism was negatively associated with the bacterial community ( β = −0.56). Conclusions These findings identify amendment-induced changes in soil nutrient status as the primary correlate of rhizosphere bacterial reorganization, with root metabolic reprogramming as a complementary plant response.

Dissolved organic matter (DOM) plays a crucial role in soil carbon sequestration, yet the molecular mechanisms by which nutrient enrichment influences DOM transformation remain poorly understood. Here, we employed high-resolution mass spectrometry and high-throughput sequencing to examine the effects of nutrient gradients on DOM transformation, defined as pairwise biochemical conversions among DOM molecules, and the associated bacterial community in green manure-amended paddy soils. Our results showcased that nutrient addition increased DOM content by 7.1%–24.2% compared to unfertilized control. Nutrient enrichment enhances DOM stability by shifting its molecular composition (toward more recalcitrant condensed aromatic- and tannin-like compounds and less lignin-like compounds) and reducing molecular diversity (i.e., the number of molecular formulas). These molecular compositional changes were accompanied by a decline in DOM transformation potential (i.e., lower reaction levels; 1.1–1.5 fold decrease) and by shifts in DOM-bacteria association patterns. Importantly, soil pH was significantly associated with these patterns, suggesting that nutrient-induced acidification may contribute to reduced DOM transformation potential. Our findings suggest a coupled nutrient–pH–microbe response that offers insights into optimizing nutrient management strategies to balance soil carbon sequestration and ecosystem resilience in paddy soils.

Bioavailable carbon (C) is a key determinant of free-living nitrogen (N) fixation, yet the regulatory role of dissolved organic matter (DOM) in paddy soils remains unclear. Using 15N2 isotope tracing, metatranscriptomics, and high-resolution mass spectrometry, we showed that phenology-dependent DOM transformations controlled rhizosphere N2 fixation throughout rice growth. N2 fixation varied significantly across phenological stages, peaking at the seedling stage (0.33 f 0.13 nmol N g- 1 h- 1) and decreasing to 0.05 f 0.02, 0.07 f 0.01, 0.13 f 0.08, and 0.14 f 0.03 nmol N g- 1 h- 1 at the tillering, heading, maturing, and harvesting stages, respectively. These changes were significantly associated with nifA transcription. Seasonal shifts in DOM toward lower combustion enthalpy and higher carbon oxidation state suggested the accumulation of more recalcitrant and energy-poor molecules. Notably, tannin-like compounds were identified as important factors influencing N2 fixation rates and nif gene expression. A potential association between methane (CH4) transformation and N2 fixation was also observed. The increase in N2 fixation rates after CH4 addition in straw-amended soils suggested that straw-derived oxidized DOM may promote diazotrophic activity under elevated CH4 conditions. Together, these results highlight the important roles of DOM dynamics and CH4 metabolism in sustaining free-living N2 fixation and provide mechanistic insights into balancing rice productivity with environmental sustainability.

Organic amendments improve soil physicochemical and microbial properties, but the effects vary by fertilizer type. These amendments also modulate the autotrophic CO₂-fixing microbial community, particularly those harboring the cbbL gene, which encodes the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) form I. Nevertheless, how cbbL-harboring autotrophs respond to different organic amendments and their associations with soil enzyme activities are still not well understood. A long-term organic amendment experiment was established in a double-cropping rice paddy field in Southern China, including four treatments: without organic fertilizer input (control), green manure (GM), pig manure (PM), and rice straw returning (RS). Soil C-, N-, and P-acquisition enzyme activities were analyzed using a fluorometric method. The cbbL-harboring bacterial community was characterized by quantitative PCR (qPCR) and high-throughput sequencing. Partial least squares path modeling (PLS-PM) was used to determine the relationships among physicochemical properties, enzyme activities, and the cbbL-harboring community. The organic amendments improved soil physicochemical properties, including pH and soil organic C (SOC). Soil C-, N-, and P- acquisition enzyme activities responded variably to the amendments. Although the cbbL gene number did not significantly change, all organic amendments reduced the diversity of cbbL-harboring bacterial community. Shifts in the cbbL-harboring community composition were also observed: GM enriched Afipia, PM favored Pseudonocardia, and RS exhibited increased abundances of Methylotenera and Sulfuricaulis. PLS-PM indicated that soil pH, SOC, and C- and N-acquisition enzyme activities negatively influenced the diversity and the composition of the cbbL-harboring community, whereas P-acquisition enzyme activity had a positive effect on the community diversity. Our study highlights the complex interactions among soil physicochemical properties, enzyme activities, and cbbL-harboring bacterial community under organic amendments. The results address the critical factors shaping the cbbL-harboring bacterial community, advancing our understanding of CO₂-fixing microorganisms in agricultural ecosystems.

This study investigated the effects of dissolved organic matter (DOM) chemical characteristics on microbial community composition under different long-term fertilization regimes in subtropical paddy soils. DOM chemical characteristics were analyzed using three-dimensional excitation-emission matrix fluorescence spectroscopy (3DEEM) combined with parallel factor analysis (PARAFAC). Microbial communities were characterized by phospholipid fatty acid (PLFA) analysis, and structural equation modeling (SEM) was applied to evaluate key drivers of microbial properties. Three fluorescence components were identified: aromatic protein and soluble microbial by-product-like substances (C1), humic acid-like substance (C2), and fulvic acid-like and humic acid-like substances (C3). Compared to the CK (no fertilizer), the NPKM (chemical nitrogen, phosphorus, and potassium fertilizer combined with organic manure) significantly increased the dissolved organic carbon (DOC) content (48.12 mg kg(-)& sup1;; p < 0.05) and the fluorescence intensities of C1-C3. PLFA analysis revealed a 71.32% increase in total fatty acid methyl esters (Total FAMEs) compared with CK, and principle component analysis (PCA) confirmed distinct shifts in microbial community structure. The humification index (HIX) was the primary factor driving microbial biomass, explaining more than 35% of the variation, while SEM indicated that DOM quantity negatively affected microbial community as represented by the PCA scores. Combined organic-inorganic fertilization enhances DOM humification and microbial activity: DOC quantity suppresses microbial community (r = -0.431) via R-strategist microbes, while DOM component diversity improves community structure (r = 0.405), potentially enhancing subtropical paddy soil fertility.

Understanding the interactions between microbial communities and soil organic matter (SOM) chemodiversity is critical for comprehending the soil carbon cycle. In this study, amplicon sequencing and ultrahigh-resolution Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR-MS) were employed to characterize soil microbial communities and SOM chemodiversity under the long-term application of different vermicompost substitution ratios for chemical fertilizers. As the vermicompost substitution ratio increased, SOM chemodiversity and microbial diversity increased, and SOM-microbial interactions tended to become more complex. Our results suggested that vermicompost application indirectly stimulated SOM chemodiversity by mediating soil microbial communities, which ultimately regulated the release of accumulated soil carbon. Notably, vermicompost application enhanced soil organic carbon mineralization, whereas the 50% vermicompost substitution ratio resulted in the lowest cumulative CO2-C release (12.16 mg CO2-C g−1 soil organic carbon) among all vermicompost treatments. Concurrently, soil organic matter content increased from 2.33% to 3.98% as the vermicompost substitution ratio increased. Therefore, a balanced substitution ratio of vermicompost for chemical fertilizers may enhance soil fertility while minimizing short-term carbon loss.

Despite the recognized importance of dissolved organic matter (DOM) in soil carbon cycling, the molecular complexity and pronounced vertical heterogeneity of DOM have hindered a mechanistic understanding of how depth-structured DOM is associated with bacterial community organization in paddy soils. Here, we combined high-resolution mass spectrometry and 16S rRNA gene sequencing to investigate linkages between DOM molecular traits and bacterial composition across soil layers (0–20, 20–40, and 40–60 cm) in four representative paddy sites in South China. DOM molecular numbers decreased markedly with depth (on average from 2,848 formulas at 0–20 cm to 1,854 formulas at 40–60 cm), accompanied by a consistent decline in DOM chemodiversity (from 7.13 to 7.08). The middle soil layer functioned as a chemical transition zone, where intensified sulfur-associated transformations enhanced DOM condensation and aromaticity. These molecular changes coincided with decreased bacterial Shannon and Chao1 indies, reduced co-occurrence network connectivity (average degree decreased from 30.83 to 24.44), and a shift from oxidative metabolism toward fermentation-dominated functions. Random forest analysis identified lipid- and protein/amino sugar-like compounds as key predictors of bacterial diversity. However, their enrichment in deeper soil layers reflected the accumulation of refractory compounds with limited microbial accessibility rather than an increase in bioavailable substrates, thereby constraining microbial niche differentiation. These findings provide insight into how depth-structured DOM covaries with microbial ecology in paddy soils and offer a foundation for improving predictions of soil carbon cycling under long-term agricultural management.

Dissolved organic matter (DOM) plays a critical role in soil carbon cycling, yet its molecular dynamics and the associated microbial mechanisms in paddy fields remain poorly understood. Here, we combined FT-ICR MS, 16S rRNA sequencing, and metatranscriptomics across rice phenological stages (seedling, tillering, heading, maturity, and harvest) to trace DOM molecular succession and reveal directional couplings between microbial activity and DOM transformation. Our results revealed stage-dependent shifts in DOM composition, with labile and oxygen-rich compounds prevailing during early rice growth, followed by the accumulation in later stages. A total of 38,420 potential transformations were detected across rice phenological stages, over 55% of which were thermodynamically limited processes, indicating that DOM succession was predominantly influenced by microbial regulation. Changes in DOM molecular composition coincided with shifts in microbial community structure, with the relative abundance of core taxa such as Thermoleophilia and Actinobacteria co-varying across rice phenological stages. Granger causality analysis suggested a directional shift from labile compounds toward aromatic and stable forms. Metatranscriptomics further showed that early expression of energy- and amino acid metabolism facilitated turnover of labile DOM, while later enrichment of lipid metabolism and auxiliary activities supported the processing of complex substrates. Together, these findings highlight stage-dependent associations between microbial dynamics and DOM molecular succession in paddy soils, providing mechanistic insights into how microbial activity influences DOM transformation processes with implications for soil carbon persistence in agroecosystems.

Understanding the effects of bio-antimicrobial materials on plant growth and against diseases and the relevant mechanisms are highly important for sustainable soil use and plant safety production. This study explored the impacts and corresponding mechanisms of the combined utilization of humic acid, chitosan, and Bacillus subtilis (B. subtilis) on tomato growth and diseases occurrence through a greenhouse pot experiment. The plant height, fresh weight, disease index, rhizosphere microbial community, and root exudates composition of tomatoes were determined. With the combined application of humic acid, chitosan and B. subtilis (HBC), the height and fresh weight of tomato plants were significantly increased (p < 0.05), and the incidence of plant diseases was decreased by 45.1%. In HBC treatment, the diversity of fungal and bacterial communities was notably enhanced. The relative abundances of Bacillus, Gemmatimonas, Neobacillus, Acinetobacter, Humicola increased, while the relative abundances of Sphingomonas, especially soil-borne plant pathogen Fusarium and Ralstonia, significantly decreased (p < 0.05). Besides the increased diversity of root exudates, the content of phenolic acids, which are allelochemicals related to continuous cropping disorder, decreased. The results of cooccurrence network analysis indicated that the abundances of Eicosanoids, Fatty acids and conjugates, and Flavonoid lycosides compounds in root exudates, which are positively correlated with pathogenic bacteria, decreased in HBC treatment. Results indicated HBC’s synergistic effect on tomato growth and disease resistance is related to its regulation of microbial community and root exudates. The study results promote the development of biological control technology and highlight its promising application in plant safety production.

The persistence of soil organic matter (SOM) is shaped by its molecular features and stability, but the temporal dynamics of these features remain unclear. Here we investigate the molecular diversity (the number of molecules) and molecular thermodynamic stability (the theoretical Gibbs free energy for the half reaction of carbon oxidation) of SOM in soils from long-term (>30 years) paddy and upland experimental fields. Thermogravimetric analysis shows that enhanced SOM thermostability aligns with the temporal variation of molecular thermodynamic stability in these soils. Increased SOM molecular thermodynamic stability occurs alongside decreased molecular diversity over decades, and this temporal trade-off (negative relationship) is modulated by increased bacterial richness. These findings highlight the role of microbial diversity in enhancing SOM thermostability and support strategies that promote bacterial richness for improved SOM persistence in agriculture. Soil organic matter stability is critical for long-term soil health and carbon sequestration. This study reveals that increased bacterial richness enhances soil organic matter thermostability by driving a trade-off between molecular diversity and thermodynamic stability.

Soil microbes play a significant role in the carbon cycle. However, our understanding of how soil microbes respond to different carbon substrates is still limited. Here, we investigated the ecological mechanisms behind the overall carbon metabolism differences under different fertilizations in a paddy ecosystem. Four fertilization treatments, no fertilizers (CK), mineral nitrogen, phosphorus and potassium fertilizers (NPK), mineral fertilizers plus organic manures (NPKM) and mineral fertilizers plus straw return (NPKS) were set up. Our results indicated that fertilization drove shifts in microbial community structure, with a reduction in the abundance of Actinobacteriota and an increase in the abundance of Chloroflexi . NPKS and CK exhibited higher carbon utilization capacity across various carbon sources, with particularly higher metabolic activity for carbohydrates than NPK and NPKM treatments. The weighted gene co‐expression network analysis (WGCNA) was used to evaluate the correlation between the modules of WGCNA and carbon metabolism. We found that microbes in the modules are important contributors to the variations of carbon metabolism. It was found that a key species in the module affecting carbohydrate utilization is C0119 , which belongs to Ktedonobacteria in the Chloroflexi . Our results suggest that fertilization could mediate core bacterial species to affect microbial utilization of different carbon substrates and finally mediate the soil carbon metabolism function.

Carbon (C), nitrogen (N) and phosphorus (P) serve as critical limiting factors for plant–microbe colonization and growth in agroecosystems, while simultaneously functioning as key regulatory controls for soil C emissions. Here, we investigated plant–microbial nutrient limitation under two states of paddy fields (cultivation state and fallow state), assessing synchronization patterns through combined analysis of plant resorption efficiency and microbial ecoenzyme stoichiometry. We found that there was a consistent pattern of N (25.00%) and P (44.44%) limitation between plant‐microbial communities. Plant–microbial communities were mainly N–limited in the rice cultivation stage and P–limited in the fallow stage. Nutrient limitation of plant–microbial communities was regulated by soil C:N:P stoichiometry: the increase in available soil N:P led to P limitation in plant communities, which enhanced the secretion of organoheterocyclic compounds and organometallic compounds. This change increased the relative abundance of Chloroflexi and Ascomycota , thereby exacerbating microbial P limitation. Notably, the direct effect of available soil N:P on microbial N:P limitation was as important as the indirect effect through plant communities. The increased available soil C:N exacerbated microbial C limitation primarily through changing microbial composition. Overall, variations in stoichiometry of available soil nutrients drove a transition from N to P limitation in plant–microbial communities during fallow periods. We highlight that fallowing is an important compensatory process to alleviate the C and N limitations of plant–microbial communities resulting from long–term agricultural monoculture patterns. Read the free Plain Language Summary for this article on the Journal blog.

Rice seedling blight, caused by various fungi, including Fusarium oxysporum, poses a severe threat to rice production. As awareness grows regarding the environmental and safety hazards associated with the application of fungicides for managing rice seedling blight, there has been a shift in focus towards biological control agents. In this study, we isolated biocontrol bacteria from paddy fields that significantly inhibited the growth of F. oxysporum in vitro and identified the strains as Bacillus amyloliquefaciens T40 and Bacillus pumilus T208. Additionally, our findings indicated that the combined application of these Bacillus strains in soil was more effective in reducing the incidence of rice seedling blight than their individual use. Analysis of 16S and internal transcribed spacer rRNA gene sequencing data revealed that the mixture of the T40 and T208 strains exhibited the lowest average clustering coefficients, which were negatively correlated with the biomass of F. oxysporum-inoculated rice seedlings. Furthermore, this mixture led to higher stochastic assembly (average |βNTI| < 2) and reduced selection pressures on rice rhizosphere bacteria compared with individual strain applications. The mixture of the T40 and T208 strains also significantly increased the expression of defense-related genes. In conclusion, the mixture of the T40 and T208 strains effectively modulates microbial community structures, enhances microbial network stability, and boosts the resistance against rice seedling blight. Our study supports the development and utilization of biological resources for crop protection.

Root exudate-mediated microbial community assembly is critical for host health and growth. However, disease-induced variations in plant-microbe interactions remain ambiguous. Here, we explored the intrinsic distinctions and interactions between the secretion patterns and microbial community composition of diseased and healthy tomatoes. Our results showed that rhizosphere microbial communities in healthy and diseased tomatoes were dominated by potentially beneficial genera (Bacillus, Rhodanobacter, Pseudolabrys, Gemmatimonas, Dongia, and Bradyrhizobium) and putative pathogens (Ralstonia and Neocosmospora), respectively, which were correlated with differential metabolites. Further metabolite addition experiments demonstrated the differential regulation mechanism of specific metabolites on host health. Drawing inspiration from synthetic communities, we displayed a way to construct the synthetic metabolites (SynMets), and we found that the SynMets (cortisol, quercetin, pyridoxal, and levodopa), which were enriched in healthy tomatoes could resist diseases by inhibiting pathogen growth and constructing beneficial microbial communities. Conversely, the SynMets (pyridoxine, N-benzylformamide, isoquinoline, and xanthine) enriched in diseased tomatoes could result in microbial imbalances by facilitating pathogen growth, thereby causing disease occurrence and growth limitation. In total, our research indicated the importance of SynMets-mediated pathogen reproduction and microbial community assembly for plant health and lays a foundation for targeted regulation of rhizosphere microecology through synthetic metabolites.