Core microbial taxonomies that maintain high organic carbon content in upland soil.

The accumulation of soil carbon (C) is crucial for the productivity and ecological function of farmland ecosystems. The balance between microbial carbon dioxide (CO2) emission and fixation determines the sustained accumulation potential of C in soil. Microorganisms involved in this process are highly obscure, thus hindering identification and further application of microorganisms with fertile soil function. In this study, a series of typical upland farmland soils were collected from 29 regions and their microbial community structure and soil C fractions were analyzed. Additionally, the rates of CO2 emission and fixation in each soil were measured. The results showed that the correlation between soil CO2 emissions and the SOC concentration was logarithmic, while that between CO2 fixation and SOC was linear. Bacterial and fungal diversity showed an upward trend with increasing soil C, and their α diversity was significantly correlated with CO2 fixation, but not correlated with CO2 emission. Fungi were more associated with soil C than bacteria, and the strength of linkage with soil C varied among the different phyla of microorganisms. Furthermore, the core microbial taxa in soils with low, medium and high SOC levels were identified by discarding redundant amplicon sequence variants, and their community differentiation was significantly driven by soil CO2 emission and fixation based on Mantel analysis. The high abundance of Chloroflexi, Nitrospirota, Actinobacteria, and Mortierellomycota in core taxa might indicate a high level of SOC level. This study highlights that SOC fluctuations are mainly driven by the core microbial taxa, rather than all microbial taxa in the agricultural system. Our research sheds light on the targeted regulation of the soil microbial community structure in upland farmland for soil fertility enhancement.

Distinct Assembly Patterns of Soil Antibiotic Resistome Revealed by Land-Use Changes over 30 Years.

Compared with the ever-growing information about the anthropogenic discharge of nutrients, metals, and antibiotics on the disturbance of antibiotic resistance genes (ARGs), less is known about how the potential natural stressors drive the evolutionary processes of antibiotic resistance. This study examined how soil resistomes evolved and differentiated over 30 years in various land use settings with spatiotemporal homogeneity and minimal human impact. We found that the contents of soil organic carbon, nitrogen, soil microbial biomass, and bioavailable heavy metals, as well as related changes in the antibiotic resistome prevalence including diversity and abundance, declined in the order of grassland > cropland > bareland. Sixty-nine remaining ARGs and 14 mobile genetic elements (MGEs) were shared among three land uses. Multiple factors (i.e., soil properties, heavy metals, bacterial community, and MGEs) contributed to the evolutionary changes of the antibiotic resistome, wherein the resistome profile was dominantly driven by MGEs from both direct and indirect pathways, supported by a partial least-squares path model analysis. Our results suggest that pathways to mitigate ARGs in soils can coincide with land degradation processes, posing a challenge to the common goal of managing our environment sustainably.

Unveiling the culturable and non-culturable actinobacterial diversity in two macroalgae species from the northern Portuguese coast.

Actinomycetota, associated with macroalgae, remains one of the least explored marine niches. The secondary metabolism of Actinomycetota, the primary microbial source of compounds relevant to biotechnology, continues to drive research into the distribution, dynamics, and metabolome of these microorganisms. In this study, we employed a combination of traditional cultivation and metagenomic analysis to investigate the diversity of Actinomycetota in two native macroalgae species from the Portuguese coast. We obtained and taxonomically identified a collection of 380 strains, which were distributed across 12 orders, 15 families, and 25 genera affiliated with the Actinomycetia class, with Streptomyces making up approximately 60% of the composition. Metagenomic results revealed the presence of Actinomycetota in both Chondrus crispus and Codium tomentosum datasets, with relative abundances of 11% and 2%, respectively. This approach identified 12 orders, 16 families, and 17 genera affiliated with Actinomycetota, with minimal overlap with the cultivation results. Acidimicrobiales emerged as the dominant actinobacterial order in both macroalgae, although no strain affiliated with this taxonomic group was successfully isolated. Our findings suggest that macroalgae represent a hotspot for Actinomycetota. The synergistic use of both culture-dependent and independent approaches proved beneficial, enabling the identification and recovery of not only abundant but also rare taxonomic members.

Anaerobic biotransformation of hexachlorocyclohexane isomers in aqueous condition: Dual CCl isotope fractionation and impact on microbial community compositions.

Hexachlorocyclohexane (HCH) isomers are persistent organic pollutants (POPs) with high toxicity, lipid solubility, chemical stability. Despite the current ban on usage of Lindane, residual contamination cannot be ignored, and HCH are frequently detected in groundwater and threaten human health. Cultures capable of degrading α-HCH, ß-HCH, γ-HCH, and δ-HCH individually have been enriched in anoxic aqueous conditions. Compound-Specific Isotope Analysis (CSIA) was applied to examine the transformation mechanisms of different HCH isomers by the four enrichment cultures. 16S rRNA sequencing techniques were employed to examine the community composition of the enrichment cultures and detect changes in these communities resulting from adding individual HCH isomers. The results indicated that the ability of the enrichment cultures for dichloroelimination of HCH isomers was inconsistent. During dichloroelimination, different bond cleavage mode of ß- and δ-HCH led to distinct isotopic effects. HCH isomers had significant impact on the microbial community, while different microbial communities showed comparable isotopic effects during the transformation of a specific HCH isomer. In addition, bacteria in the phyla Proteobacteria and Firmicutes were proposed as the dominant dechlorinators. This study provides a novel perspective on the mode of bond cleavage during HCH dichloroelimination and the effect of HCH on microbial communities, which could potentially support the evaluation of HCH transformation by CSIA and their effects on the microecosystems of groundwater.

A novel bioprospecting strategy via 13C-based high-throughput probing of active methylotrophs inhabiting oil reservoir surface soil.

Methane-oxidizing bacteria (MOB) have long been considered as a microbial indicator for oil and gas prospecting. However, due to the phylogenetically narrow breath of ecophysiologically distinct MOB, classic culture-dependent approaches could not discriminate MOB population at fine resolution, and accurately reflect the abundance of active MOB in the soil above oil and gas reservoirs. Here, we presented a novel microbial anomaly detection (MAD) strategy to quantitatively identify specific indicator methylotrophs in the surface soils for bioprospecting oil and gas reservoirs by using a combination of 13C-DNA stable isotope probing (SIP), high-throughput sequencing (HTS), quantitative PCR (qPCR) and geostatistical analysis. The Chunguang oilfield of the Junggar Basin was selected as a model system in western China, and type I methanotrophic Methylobacter was most active in the topsoil above the productive oil wells, while type II methanotrophic Methylosinus predominated in the dry well soils, exhibiting clear differences between non- and oil reservoir soils. Similar results were observed by quantification of Methylobacter pmoA genes as a specific bioindicator for the prediction of unknown reservoirs by grid sampling. A microbial anomaly distribution map based on geostatistical analysis further showed that the anomalous zones were highly consistent with petroleum, geological and seismic data, and validated by subsequent drilling. Over seven years, a total of 24 wells have been designed and drilled into the targeted anomaly, and the success rate via the MAD prospecting strategy was 83 %. Our results suggested that molecular techniques are powerful tools for oil and gas prospecting. This study indicates that the exploration efficiency could be significantly improved by integrating multi-disciplinary information in geophysics and geomicrobiology while reducing the drilling risk to a greater extent.

Changes in bacterial communities during rice cultivation remove phenolic constraints on peatland carbon preservation.

Northern peatlands contain ~30% of terrestrial carbon (C) stores, but in recent decades, 14% to 20% of the stored C has been lost because of conversion of the peatland to cropland. Microorganisms are widely acknowledged as primary decomposers, but the keystone taxa within the bacterial community regulating C loss from cultivated peatlands remain largely unknown. In this study, we investigated the bacterial taxa driving peat C mineralization during rice cultivation. Cultivation significantly decreased concentrations of soil organic C, dissolved organic C (DOC), carbohydrates, and phenolics but increased C mineralization rate (CMR). Consistent with the classic theory that phenolic inhibition creates a "latch" that reduces peat C decomposition, phenolics were highly negatively correlated with CMR in cultivated peatlands, indicating that elimination of inhibitory phenolics can accelerate soil C mineralization. Bacterial communities were significantly different following peatland cultivation, and co-occurrence diagnosis analysis revealed substantial changes in network clusters of closely connected nodes (modules) and bacterial keystone taxa. Specifically, in cultivated peatlands, bacterial modules were significantly negatively correlated with phenolics, carbohydrates, and DOC. While keystone taxa Xanthomonadales, Arthrobacter, and Bacteroidetes_vadinHA17 can regulate bacterial modules and promote carbon mineralization. Those observations indicated that changes in bacterial modules can promote phenolic decomposition and eliminate phenolic inhibition of labile C decomposition, thus accelerating soil organic C loss during rice cultivation. Overall, the study provides deeper insights into microbe-driven peat C loss during rice cultivation and highlights the crucial role of keystone bacterial taxa in the removal of phenolic constraints on peat C preservation.

Ridge with no-tillage facilitates microbial N2 fixation associated with methane oxidation in rice soil.

Aerobic methane-oxidizing bacteria (MOB) play a crucial role in mitigating the greenhouse gas methane emission, particularly prevalent in flooded wetlands. The implementation of ridge with no-tillage practices within a rice-rape rotation system proves effective in overcoming the restrictive redox conditions associated with waterlogging. This approach enhances capillary water availability from furrows, especially during periods of low rainfall, thereby supporting plant growth on the ridges. However, the microbe-mediated accumulation of soil organic carbon and nitrogen remains insufficiently understood under this agricultural practice, particularly concerning methane oxidation, which holds ecological and agricultural significance in the rice fields. In this study, the ridge and ditch soils from a 28-year-old ridge with no-tillage rice field experiment were utilized for incubation with 13C-CH4 and 15NN2 to estimate the methane-oxidizing and N2-fixing potentials. Our findings reveal a significantly higher net production of fresh soil organic carbon in the ridge compared to the ditch soil during methane oxidation, with values of 626 and 543 µg 13C g-1 dry weight soil, respectively. Additionally, the fixed 15N exhibited a twofold increase in the ridge soil (14.1 µg 15N g-1 dry weight soil) compared to the ditch soil. Interestingly, the result of DNA-based stable isotope probing indicated no significant differences in active MOB and N2 fixers between ridge and ditch soils. Both Methylocystis-like type II and Methylosarcina/Methylomonas-like type I MOB catalyzed methane into organic biomass carbon pools. Soil N2-fixing activity was associated with the 15N-labeling of methane oxidizers and non-MOB, such as methanol oxidizers (Hyphomicrobium) and conventional N2 fixers (Burkholderia). Methane oxidation also fostered microbial interactions, as evidenced by co-occurrence patterns. These results underscore the dual role of microbial methane oxidation - not only as a recognized sink for the potent greenhouse gas methane but also as a source of soil organic carbon and bioavailable nitrogen. This emphasizes the pivotal role of microbial methane metabolism in contributing to soil carbon and nitrogen accumulation in ridge with no-tillage systems.

Identifying Active Rather than Total Methanotrophs Inhabiting Surface Soil Is Essential for the Microbial Prospection of Gas Reservoirs.

Methane-oxidizing bacteria (MOB) have long been recognized as an important bioindicator for oil and gas exploration. However, due to their physiological and ecological diversity, the distribution of MOB in different habitats varies widely, making it challenging to authentically reflect the abundance of active MOB in the soil above oil and gas reservoirs using conventional methods. Here, we selected the Puguang gas field of the Sichuan Basin in Southwest China as a model system to study the ecological characteristics of methanotrophs using culture-independent molecular techniques. Initially, by comparing the abundance of the pmoA genes determined by quantitative PCR (qPCR), no significant difference was found between gas well and non-gas well soils, indicating that the abundance of total MOB may not necessarily reflect the distribution of the underlying gas reservoirs. 13C-DNA stable isotope probing (DNA-SIP) in combination with high-throughput sequencing (HTS) furthermore revealed that type II methanotrophic Methylocystis was the absolutely predominant active MOB in the non-gas-field soils, whereas the niche vacated by Methylocystis was gradually filled with type I RPC-2 (rice paddy cluster-2) and Methylosarcina in the surface soils of gas reservoirs after geoscale acclimation to trace- and continuous-methane supply. The sum of the relative abundance of RPC-2 and Methylosarcina was then used as specific biotic index (BI) in the Puguang gas field. A microbial anomaly distribution map based on the BI values showed that the anomalous zones were highly consistent with geological and geophysical data, and known drilling results. Therefore, the active but not total methanotrophs successfully reflected the microseepage intensity of the underlying active hydrocarbon system, and can be used as an essential quantitative index to determine the existence and distribution of reservoirs. Our results suggest that molecular microbial techniques are powerful tools for oil and gas prospecting.

Intensive N2 fixation accelerates microbial turnover in cropland soils.

Biological nitrogen fixation (BNF) is strongly affected by the carbon (C) and nitrogen (N) stoichiometry in soil and depends on the input of organic C. Due to the high metabolic costs of nitrogenase activity, however, the response of BNF to organic C input and its impact on microbial turnover remain unclear. To address this knowledge gap, we combined 15N2 tracing with high-throughput sequencing by adding glucose or glucose plus mineral N fertilizer for a 12-day incubation in three cropland soils. Glucose addition alone strongly changed the BNF activity (0.76-2.51 mg N kg-1 d-1), while BNF was completely absent after mineral N fertilization. This switch-on of BNF by glucose addition supported equally high rates of microbial growth and organic C mineralization compared with the direct mineral N assimilation by microorganisms. Glucose-induced BNF was predominantly catalyzed by Azotobacter-affiliated free-living diazotrophs (>50 % of the total nifH genes), which increased with diverse nondiazotrophs such as Nitrososphaera, Bacillus and Pseudoxanthomonas. Structural equation models (SEMs) and random forest (RF) analyses consistently revealed that the soil C:N ratio and Azotobacter-affiliated diazotrophic abundances were the key factors affecting glucose-induced BNF. Our findings emphasize the importance of free-living diazotrophs for microbial turnover of organic C in soil.

Long-term conservation tillage enhances microbial carbon use efficiency by altering multitrophic interactions in soil.

Microbial carbon (C) use efficiency (CUE) plays a key role in soil C storage. The predation of protists on bacteria and fungi has potential impacts on the global C cycle. However, under conservation tillage conditions, the effects of multitrophic interactions on soil microbial CUE are still unclear. Here, we investigate the multitrophic network (especially the keystone ecological cluster) and its regulation of soil microbial CUE and soil organic C (SOC) under different long-term (15-year) tillage practices. We found that conservation tillage (CT) significantly enhanced microbial CUE, turnover, and SOC (P < 0.05) compared to traditional tillage (control, CK). At the same time, tillage practice and soil depth had significant effects on the structure of fungal and protistan communities. Furthermore, the soil biodiversity of the keystone cluster was positively correlated with the microbial physiological traits (CUE, microbial growth rate (MGR), microbial respiration rate (Rs), microbial turnover) and SOC (P < 0.05). Protistan richness played the strongest role in directly shaping the keystone cluster. Compared with CK, CT generally enhanced the correlation between microbial communities and microbial physiological characteristics and SOC. Overall, our results illustrate that the top-down control (the organisms at higher trophic levels affect the organisms at lower trophic levels) of protists in the soil micro-food web plays an important role in improving microbial CUE under conservation tillage. Our findings provide a theoretical basis for promoting the application of protists in targeted microbial engineering and contribute to the promotion of conservation agriculture and the improvement of soil C sequestration potential.

Amendment-driven soil health restoration through soil pH and microbial robustness in a Cd/Cu-combined acidic soil: A ten-year in-situ field experiment.

Soil health arguably depends on biodiversity and has received wide attention in heavy-metal (HM) contaminated farmland remediation in recent years. However, long-term effects and mechanisms of soil amendment remain poorly understood with respect to soil microbal community. In this in-situ field study, four soil amendments (attapulgite-At, apatite-Ap, montmorillonite-M, lime-L) at three rates were applied once only for ten years in a cadmium (Cd)-copper (Cu) contaminated paddy soil deprecated for over five years. Results showed that after ten years and in compared with CK (no amendment), total Cd concentration and its risk in plot soils were not altered by amendments (p > 0.05), but total Cu concentration and its risk were significantly increased by both Ap and L, especially the former, rather than At and M (p < 0.05), through increased soil pH and enhanced bacterial alpha diversity as well as plant community. Soil microbial communities were more affected by amendment type (30%) than dosage (11%), microbial network characteristics were dominated by rare taxa, and soil multifunctionality was improved in Ap- and L-amended soils. A structural equation model (SEM) indicated that 57.3% of soil multifunctionality variances were accounted for by soil pH (+0.696) and microbial network robustness (-0.301). Moreover, microbial robustness could be potentially used as an indicator of soil multifunctionality, and Ap could be optimized to improve soil health in combined with biomass removal. These findings would advance the understanding of soil microbial roles, especially its network robustness, on soil multifunctionality for the remediation of metal contaminated soils and metal control management strategies in acidic soils. ENVIRONMENTAL IMPLICATION: Farmland soil contamination by heavy metals (HMs) has been becoming a serious global environmental challenge. However, most studies have been conducted over the short term, leading to a gap in the long-term remediation efficiency and ecological benefits of soil amendments. For the successful deployment of immobilization technologies, it is critical to understand the long-term stability of the immobilized HMs and soil health. Our study, to the best of our knowlege, is the first to state the long-term effects and mechanisms of soil amendments on soil health and optimize an effective and eco-friendly amendment for long-term Cd/Cu immobilization.

Occurrence, distribution and ecological risk assessment of herbicide residues in cropland soils from the Mollisols region of Northeast China.

The first systematic and comprehensive investigation of herbicide residues was conducted by identifying their spatial distribution, influencing factors and ecological risk in cropland soils from the Mollisols region covering 109 million hm2 in Northeast China. Fifty-six herbicides were detected with total herbicide concentrations ranging from 1.01 to 1558.13 µg/kg (mean: 227.45). Atrazine, its degradates deethyl atrazine (DEA) and deisopropyl atrazine (DIA), trifluralin and butachlor were the most frequently detected herbicides, while DEA, clomazone, nicosulfuron, fomesafen, and mefenacet exhibited the highest concentrations. Despite being less frequently reported in Chinese soils, fomesafen, nicosulfuron, clomazone, and mefenacet were found widely present. Although most of the compounds posed a minimal or low ecological risk, atrazine, nicosulfuron and DEA exhibited medium to high potential risks. The key factors identified to regulate the fate of herbicides were soil chemical properties, amount of herbicides application, and the crop type. The soybean soils showed highest herbicide residues, while the soil mineral contents likely adsorbed more herbicides. This study provides a valuable large-scale dataset of herbicide residues across the entire Mollisols region of China along with fine-scale characterization of the ecological risks. Mitigation and management measures are needed to reduce the herbicide inputs and residues in the region.

Parabacteroides distasonis uses dietary inulin to suppress NASH via its metabolite pentadecanoic acid.

Non-alcoholic steatohepatitis (NASH) is the severe form of non-alcoholic fatty liver disease, and is characterized by liver inflammation and fat accumulation. Dietary interventions, such as fibre, have been shown to alleviate this metabolic disorder in mice via the gut microbiota. Here, we investigated the mechanistic role of the gut microbiota in ameliorating NASH via dietary fibre in mice. Soluble fibre inulin was found to be more effective than insoluble fibre cellulose to suppress NASH progression in mice, as shown by reduced hepatic steatosis, necro-inflammation, ballooning and fibrosis. We employed stable isotope probing to trace the incorporation of 13C-inulin into gut bacterial genomes and metabolites during NASH progression. Shotgun metagenome sequencing revealed that the commensal Parabacteroides distasonis was enriched by 13C-inulin. Integration of 13C-inulin metagenomes and metabolomes suggested that P. distasonis used inulin to produce pentadecanoic acid, an odd-chain fatty acid, which was confirmed in vitro and in germ-free mice. P. distasonis or pentadecanoic acid was protective against NASH in mice. Mechanistically, inulin, P. distasonis or pentadecanoic acid restored gut barrier function in NASH models, which reduced serum lipopolysaccharide and liver pro-inflammatory cytokine expression. Overall this shows that gut microbiota members can use dietary fibre to generate beneficial metabolites to suppress metabolic disease.

Combined Effects of Temperature and Dietary Lipid Level on Body Composition, Growth, and Freshness Profile in European Seabass, Dicentrarchus labrax.

A fish trial was carried out to evaluate the combined effects of temperature and dietary lipid level on the body composition, growth performance, and freshness profile of the European seabass (Dicentrarchus labrax). Fish were kept for 56 days at 20 °C and 24 °C and fed on two diets, with 16% and 20% lipid. At the end of the trial, fish were euthanized at two temperature conditions (0.6 °C or -0.6 °C) and kept on ice for 10 days at 4 °C to evaluate their freshness condition. Findings demonstrated that fish reared at 24 °C presented a lower lipid level and a higher daily growth index than those at 20 °C. Additionally, sensory analysis (Quality Index Method-QIM) and microbiological analysis revealed that fish reared at 24 °C showed better freshness conditions than those at 20 °C. However, the 16S rRNA metabarcoding analyses revealed a higher proliferation of genera associated with fish-spoiling bacteria in the skin microbiome of fish reared at 24 °C, i.e., Vibrio and Acinetobacter, which was not observed in the skin microbiome of fish reared at 20 °C. Nevertheless, the dietary lipid level did not have any influence on fish freshness. Therefore, our data suggest that the increase in temperature to 24 °C is beneficial for the growth and freshness profile (lower QIM and lower CFUs/cm2) of this particular species. Additionally, the lower euthanasia temperature (-0.6 °C) seems to lead to higher fish freshness than the normal temperature (0.6 °C).

15N tracing reveals preference for different nitrogen forms of Fusarium oxysporum f. sp. cubense tropical race 4.

Plant uptake of nitrogen is often associated with increased incidence of banana Fusarium wilt, a disease caused by the soil-borne fungus Fusarium oxysporum f. sp. cubense tropical race 4 (Foc TR4). However, the nitrogen metabolic preferences of Foc TR4 pathogens remain unknown. In this study, we investigated the ecophysiological patterns of Foc TR4 grown on different combinations of organic and inorganic nitrogen. Potato Dextrose Agar (PDA) and Rose Bengal Medium (RBM) were used as an organic nitrogen source, which was sequentially replaced with inorganic N (0, 50% or 90%) in the form 15NH4NO3 or NH4 15NO3 to reveal preferential assimilation of ammonium or nitrate. The results showed that mycelium biomass and nitrogen content decreased significantly, while the carbon content and C:N ratio increased in Foc TR4 grown on media containing inorganic nitrogen sources. Mycelium biomass was negatively correlated with C:N ratio. Mycelium 15N abundance increased significantly between the PDA50 + A50/RBM50 + A50 treatments (50% organic nitrogen+50%15NH4NO3) and the PDA10 + A90/RBM10 + A90 treatments (10% organic nitrogen+90%15NH4NO3). These results indicate that the higher C:N ratio reduced mycelium growth by reducing its biomass and diameter and showed that Foc TR4 preferred to use ammonium nitrogen to promote the growth. These findings suggest that treating banana crops with a combination of organic and inorganic (i.e., nitrate) nitrogen could be a better way to defend against Fusarium wilt of banana.

Selection of rhizosphere communities of diverse rotation crops reveals unique core microbiome associated with reduced banana Fusarium wilt disease.

Crop rotation can assemble distinct core microbiota as functionally specific barriers against the invasion of banana Fusarium oxysporum pathogens. However, the taxonomic identity of rotation-unique core taxa and their legacy effects are poorly understood under field conditions. Pepper and eggplant rotations were employed to reveal rotation crop- and banana-unique antagonistic core taxa by in situ tracking of the soil microbiome assembly patterns for 2 yr. The rotation crop-unique antagonistic taxa were isolated and functionally verified by culture-dependent techniques, high-throughput sequencing, and pot experiments. Pepper and eggplant rotations resulted in eight and one rotation-unique antagonistic core taxa out of 12 507 microbial taxa, respectively. These nine antagonistic taxa were retained the following year and significantly decreased banana wilt disease incidence via legacy effects, although the cultivated strains were exclusively of the genera Bacillus and Pseudomonas. The fermentation broth and volatiles of these two taxa showed strong antagonistic activity, and pot experiments demonstrated high suppression of wilt disease and significant promotion of banana growth. Our study provides a mechanistic understanding of the identification of rotation crop-unique antagonistic taxa and highlights the importance of targeted cultivation of beneficial microorganisms for optimizing crop rotation-based scenarios in support of banana agriculture sustainability.

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

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.

Methanotrophy Alleviates Nitrogen Constraint of Carbon Turnover by Rice Root-Associated Microbiomes.

The bioavailability of nitrogen constrains primary productivity, and ecosystem stoichiometry implies stimulation of N2 fixation in association with carbon sequestration in hotspots such as paddy soils. In this study, we show that N2 fixation was triggered by methane oxidation and the methanotrophs serve as microbial engines driving the turnover of carbon and nitrogen in rice roots. 15N2-stable isotope probing showed that N2-fixing activity was stimulated 160-fold by CH4 oxidation from 0.27 to 43.3 µmol N g-1 dry weight root biomass, and approximately 42.5% of the fixed N existed in the form of 15N-NH4 + through microbial mineralization. Nitrate amendment almost completely abolished N2 fixation. Ecophysiology flux measurement indicated that methane oxidation-induced N2 fixation contributed only 1.9% of total nitrogen, whereas methanotrophy-primed mineralization accounted for 21.7% of total nitrogen to facilitate root carbon turnover. DNA-based stable isotope probing further indicated that gammaproteobacterial Methylomonas-like methanotrophs dominated N2 fixation in CH4-consuming roots, whereas nitrate addition resulted in the shift of the active population to alphaproteobacterial Methylocystis-like methanotrophs. Co-occurring pattern analysis of active microbial community further suggested that a number of keystone taxa could have played a major role in nitrogen acquisition through root decomposition and N2 fixation to facilitate nutrient cycling while maintaining soil productivity. This study thus highlights the importance of root-associated methanotrophs as both biofilters of greenhouse gas methane and microbial engines of bioavailable nitrogen for rice growth.

Phylogeny and Metabolic Potential of the Methanotrophic Lineage MO3 in Beijerinckiaceae from the Paddy Soil through Metagenome-Assembled Genome Reconstruction.

Although the study of aerobic methane-oxidizing bacteria (MOB, methanotrophs) has been carried out for more than a hundred years, there are many uncultivated methanotrophic lineages whose metabolism is largely unknown. Here, we reconstructed a nearly complete genome of a Beijerinckiaceae methanotroph from the enrichment of paddy soil by using nitrogen-free M2 medium. The methanotroph labeled as MO3_YZ.1 had a size of 3.83 Mb, GC content of 65.6%, and 3442 gene-coding regions. Based on phylogeny of pmoA gene and genome and the genomic average nucleotide identity, we confirmed its affiliation to the MO3 lineage and a close relationship to Methylocapsa. MO3_YZ.1 contained mxaF- and xoxF-type methanol dehydrogenase. MO3_YZ.1 used the serine cycle to assimilate carbon and regenerated glyoxylate through the glyoxylate shunt as it contained isocitrate lyase and complete tricarboxylic acid cycle-coding genes. The ethylmalonyl-CoA pathway and Calvin-Benson-Bassham cycle were incomplete in MO3_YZ.1. Three acetate utilization enzyme-coding genes were identified, suggesting its potential ability to utilize acetate. The presence of genes for N2 fixation, sulfur transformation, and poly-ß-hydroxybutyrate synthesis enable its survival in heterogeneous habitats with fluctuating supplies of carbon, nitrogen, and sulfur.