ER body-resident myrosinases and tryptophan specialized metabolism modulate root microbiota assembly.

Endoplasmic reticulum (ER) bodies are ER-derived structures that contain a large amount of PYK10 myrosinase, which hydrolyzes tryptophan (Trp)-derived indole glucosinolates (IGs). Given the well-described role of IGs in root-microbe interactions, we hypothesized that ER bodies in roots are important for interaction with soil-borne microbes at the root-soil interface. We used mutants impaired in ER bodies (nai1), ER body-resident myrosinases (pyk10bglu21), IG biosynthesis (myb34/51/122), and Trp specialized metabolism (cyp79b2b3) to profile their root microbiota community in natural soil, evaluate the impact of axenically collected root exudates on soil or synthetic microbial communities, and test their response to fungal endophytes in a mono-association setup. Tested mutants exhibited altered bacterial and fungal communities in rhizoplane and endosphere, respectively. Natural soils and bacterial synthetic communities treated with mutant root exudates exhibited distinctive microbial profiles from those treated with wild-type (WT) exudates. Most tested endophytes severely restricted the growth of cyp79b2b3, a part of which also impaired the growth of pyk10bglu21. Our results suggest that root ER bodies and their resident myrosinases modulate the profile of root-secreted metabolites and thereby influence root-microbiota interactions.

Unraveling the Roles of Neuropeptides in the Chemosensation of the Root-Knot Nematode Meloidogyne javanica.

The identification of novel drug targets in plant-parasitic nematodes (PPNs) is imperative due to the loss of traditional nematicides and a lack of replacements. Chemosensation, which is pivotal for PPNs in locating host roots, has become a focus in nematode behavioral research. However, its underlying molecular basis is still indistinct in such a diverse group of PPNs. To characterize genes participating in chemosensation in the Javanese root-knot nematode Meloidogyne javanica, RNA-sequencing of the second-stage juveniles (J2s) treated with tomato root exudate (TRE) for 1 h and 6 h was performed. Genes related to chemosensation in M. javanica mainly responded to TRE treatment at 1 h. Moreover, a gene ontology (GO) analysis underscored the significance of the neuropeptide G protein-coupled receptor signaling pathway. Consequently, the repertoire of putative neuropeptides in M. javanica, including FMRFamide-like peptides (FLPs), insulin-like peptides (ILPs), and neuropeptide-like peptides (NLPs), were outlined based on a homology analysis. The gene Mjflp-14a, harboring two neuropeptides, was significantly up-regulated at 1 h TRE treatment. Through peptide synthesis and J2 treatment, one of the two neuropeptides (MjFLP-14-2) was proven to influence the J2 chemotaxis towards tomato root tips. Overall, our study reinforces the potential of nematode neuropeptides as novel targets and tools for root-knot nematode control.

The Phosphorus-Iron Nexus: Decoding the Nutrients Interaction in Soil and Plant.

Phosphorus (P) and iron (Fe) are two essential mineral nutrients in plant growth. It is widely observed that interactions of P and Fe could influence their availability in soils and affect their homeostasis in plants, which has received significant attention in recent years. This review presents a summary of latest advances in the activation of insoluble Fe-P complexes by soil properties, microorganisms, and plants. Furthermore, we elucidate the physiological and molecular mechanisms underlying how plants adapt to Fe-P interactions. This review also discusses the current limitations and presents potential avenues for promoting sustainable agriculture through the optimization of P and Fe utilization efficiency in crops.

[Interaction between root exudates of medicinal plants and rhizosphere microorganisms and its application in ecological planting of Chinese medicinal materials].

The rhizosphere is an important place for material exchange between medicinal plants and soil. Root exudates are the medium of material and signal exchange between plants and soil and are the key factors in the regulation of rhizosphere microecology. Rhizosphere microorganisms are an important part of the rhizosphere microecology of medicinal plants, and the interaction between root exudates and rhizosphere microorganisms has an important influence on the growth and quality formation of medicinal plants. Rational utilization of the interaction between root exudates and rhizosphere microorganisms of medicinal plants is one of the important ways to ensure the healthy growth of medicinal plants and promote the development of ecological planting of Chinese medicinal materials. In the paper, the research status of root exudates and rhizosphere microorganisms of medicinal plants in recent years was summarized. The interaction mechanism between root exudates and rhizosphere microorganisms of medicinal plants, as well as the influence of rhizosphere microorganisms on the growth of medicinal plants, were analyzed. In addition, the advantages and promoting effects of intercropping ecological planting mode on rhizosphere microecology of medicinal plants and quality improvement of Chinese medicinal materials were explained, providing a good basis for the study of the interaction among medicinal plants, microorganisms, and soil. Furthermore, it could produce important theoretical and practical significance for the ecological planting and sustainable utilization of medicinal plants.

Extended automated quantification algorithm (AQuA) for targeted 1H NMR metabolomics of highly complex samples: application to plant root exudates.

INTRODUCTION: The Automated Quantification Algorithm (AQuA) is a rapid and efficient method for targeted NMR-based metabolomics, currently optimised for blood plasma. AQuA quantifies metabolites from 1D-1H NMR spectra based on the height of only one signal per metabolite, which minimises the computational time and workload of the method without compromising the quantification accuracy. OBJECTIVES: To develop a fast and computationally efficient extension of AQuA for quantification of selected metabolites in highly complex samples, with minimal prior sample preparation. In particular, the method should be capable of handling interferences caused by broad background signals. METHODS: An automatic baseline correction function was combined with AQuA into an automated workflow, the extended AQuA, for quantification of metabolites in plant root exudate NMR spectra that contained broad background signals and baseline distortions. The approach was evaluated using simulations as well as a spike-in experiment in which known metabolite amounts were added to a complex sample matrix. RESULTS: The extended AQuA enables accurate quantification of metabolites in 1D-1H NMR spectra with varying complexity. The method is very fast (< 1 s per spectrum) and can be fully automated. CONCLUSIONS: The extended AQuA is an automated quantification method intended for 1D-1H NMR spectra containing broad background signals and baseline distortions. Although the method was developed for plant root exudates, it should be readily applicable to any NMR spectra displaying similar issues as it is purely computational and applied to NMR spectra post-acquisition.

Meloidogyne enterolobii-induced Changes in Guava Root Exudates Are Associated With Root Rotting Caused by Neocosmospora falciformis.

Despite the worldwide importance of disease complexes involving root-feeding nematodes and soilborne fungi, there have been few in-depth studies on how these organisms interact at the molecular level. Previous studies of guava decline have shown that root exudates from Meloidogyne enterolobii-parasitized guava plants (NP plants), but not from nematode-free plants (NF plants), enable the fungus Neocosmospora falciformis to rot guava roots, leading to plant death. To further characterize this interaction, NP and NF root exudates were lyophilized; extracted with distinct solvents; quantified regarding amino acids, soluble carbohydrates, sucrose, phenols, and alkaloids; and submitted to a bioassay to determine their ability to enable N. falciformis to rot the guava seedlings' roots. NP root exudates were richer than NF root exudates in amino acids, carbohydrates, and sucrose. Only the fractions NP-03 and NP-04 enabled fungal root rotting. NP-03 was then sequentially fractionated through chromatographic silica columns. At each step, the main fractions were reassessed in bioassay. The final fraction that enabled fungal root rotting was submitted to analysis using high performance liquid chromatography, nuclear magnetic resonance, mass spectrometry, energy-dispersive X-ray fluorescence, and computational calculations, leading to the identification of 1,5-dinitrobiuret as the predominant substance. In conclusion, parasitism by M. enterolobii causes an enrichment of guava root exudates that likely favors microorganisms capable of producing 1,5-dinitrobiuret in the rhizosphere. The accumulation of biuret, a known phytotoxic substance, possibly hampers root physiology and the innate immunity of guava to N. falciformis.

Carboxylic acid accumulation and secretion contribute to the alkali-stress tolerance of halophyte Leymus chinensis.

Leymus chinensis is a dominant halophytic grass in alkalized grasslands of Northeast China. To explore the alkali-tolerance mechanism of L. chinensis, we applied a widely targeted metabolomic approach to analyze metabolic responses of its root exudates, root tissues and leaves under alkali-stress conditions. L. chinensis extensively secreted organic acids, phenolic acids, free fatty acids and other substances having -COOH or phosphate groups when grown under alkali-stress conditions. The buffering capacity of these secreted substances promoted pH regulation in the rhizosphere during responses to alkali stress. L. chinensis leaves exhibited enhanced accumulations of free fatty acids, lipids, amino acids, organic acids, phenolic acids and alkaloids, which play important roles in maintaining cell membrane stability, regulating osmotic pressure and providing substrates for the alkali-stress responses of roots. The accumulations of numerous flavonoids, saccharides and alcohols were extensively enhanced in the roots of L. chinensis, but rarely enhanced in the leaves, under alkali-stress conditions. Enhanced accumulations of flavonoids, saccharides and alcohols increased the removal of reactive oxygen species and alleviated oxygen damage caused by alkali stress. In this study, we revealed the metabolic response mechanisms of L. chinensis under alkali-stress conditions, emphasizing important roles for the accumulation and secretion of organic acids, amino acids, fatty acids and other substances in alkali tolerance.

MAPK Cascades in Plant Microbiota Structure and Functioning.

Mitogen-activated protein kinase (MAPK) cascades are highly conserved signaling modules that coordinate diverse biological processes such as plant innate immunity and development. Recently, MAPK cascades have emerged as pivotal regulators of the plant holobiont, influencing the assembly of normal plant microbiota, essential for maintaining optimal plant growth and health. In this review, we provide an overview of current knowledge on MAPK cascades, from upstream perception of microbial stimuli to downstream host responses. Synthesizing recent findings, we explore the intricate connections between MAPK signaling and the assembly and functioning of plant microbiota. Additionally, the role of MAPK activation in orchestrating dynamic changes in root exudation to shape microbiota composition is discussed. Finally, our review concludes by emphasizing the necessity for more sophisticated techniques to accurately decipher the role of MAPK signaling in establishing the plant holobiont relationship.

Anthropogenic fertilization influences a shift in barley rhizosphere microbial communities.

Background: Anthropogenic mediations contribute a significant role in stimulating positive reactions in soil-plant interactions; however, methodical reports on how anthropogenic activities impact soil microorganism-induced properties and soil health are still inadequate. In this study, we evaluated the influence of anthropogenic fertilization of farmland soil on barley rhizosphere microbial community structure and diversity, and the significant impacts on agro-ecosystem productivity. This will help validate the premise that soil amendment with prolonged synthetic fertilizers can lead to a significant reduction in bacterial abundance and diversity, while soils amended with organic fertilizers elicit the succession of the native soil microbial community and favor the growth of copiotrophic bacteria. Methods: The total metagenomic DNA was extracted from soils obtained from the barley rhizosphere under chemical fertilization (CB), organic fertilization (OB), and bulk soil (NB). Subsequently, these samples were sequenced using an amplicon-based sequencing approach, and the raw sequence dataset was examined using a metagenomic rast server (MG-RAST). Results: Our findings showed that all environments (CB, OB, and NB) shared numerous soil bacterial phyla but with different compositions. However, Bacteroidetes, Proteobacteria, and Actinobacteria predominated in the barley rhizosphere under chemical fertilization, organic fertilization, and bulk soils, respectively. Alpha and beta diversity analysis showed that the diversity of bacteria under organic barley rhizosphere was significantly higher and more evenly distributed than bacteria under chemical fertilization and bulk soil. Conclusion: Understanding the impact of conventional and organic fertilizers on the structure, composition, and diversity of the rhizosphere microbiome will assist in soil engineering to enhance microbial diversity in the agroecosystem.

Polyethylene microplastic modulates lettuce root exudates and induces oxidative damage under prolonged hydroponic exposure.

Root exudates are pivotal in plant stress responses, however, the impact of microplastics (MPs) on their release and characteristics remains poorly understood. This study delves into the effects of 0.05 % and 0.1 % (w/w) additions of polyethylene (PE) MPs on the growth and physiological properties of lettuce (Lactuca sativa L.) following 28 days of exposure. The release characteristics of root exudates were assessed using UV-vis and 3D-EEM. The results indicated that PE increased leaf number but did not significantly affect other agronomic traits or pigment contents. Notably, 0.05 % PE increased the total root length and surface area compared to the 0.1 % addition, while a non-significant trend towards decreased root activity was observed with PE MPs. PE MPs with 0.1 % addition notably reduced the DOC concentration in root exudates by 37.5 %, while 0.05 % PE had no impact on DOC and DON concentrations. PE addition increased the SUVA254, SUVA260, and SUVA280 values of root exudates, with the most pronounced effect seen in the 0.05 % PE treatment. This suggests an increase of aromaticity and hydrophobic components induced by PE addition. Fluorescence Regional Integration (FRI) analysis of 3D-EEM revealed that aromatic proteins (region I and II) were dominant in root exudates, with a slight increase in fulvic acid-like substances (region III) under 0.1 % PE addition. Moreover, prolonged PE exposure induced ROS damage in lettuce leaves, evidenced by a significant increase in content and production rate of O2·-. The decrease in CAT and POD activities may account for the lettuce's response to environmental stress, potentially surpassing its tolerance threshold or undergoing adaptive regulation. These findings underscore the potential risk of prolonged exposure to PE MPs on lettuce growth.

The Function of Root Exudates in the Root Colonization by Beneficial Soil Rhizobacteria.

Soil-beneficial microbes in the rhizosphere play important roles in improving plant growth and health. Root exudates play key roles in plant-microbe interactions and rhizobacterial colonization. This review describes the factors influencing the dynamic interactions between root exudates and the soil microbiome in the rhizosphere, including plant genotype, plant development, and environmental abiotic and biotic factors. We also discuss the roles of specific metabolic mechanisms, regulators, and signals of beneficial soil bacteria in terms of colonization ability. We highlight the latest research progress on the roles of root exudates in regulating beneficial rhizobacterial colonization. Organic acids, amino acids, sugars, sugar alcohols, flavonoids, phenolic compounds, volatiles, and other secondary metabolites are discussed in detail. Finally, we propose future research objectives that will help us better understand the role of root exudates in root colonization by rhizobacteria and promote the sustainable development of agriculture and forestry.

Responses in Plant Growth and Root Exudates of Pistia stratiotes under Zn and Cu Stress.

At present, the situation regarding heavy metal pollution in aquatic environments is becoming more and more serious. The bioaccumulation of heavy metals in aquatic plants causes obvious phytotoxicity, which can also induce secondary pollution in the aquatic environment. Zinc and copper, as indispensable elements for plant growth, are also prominent heavy metals in water pollution in China, and their concentrations play a crucial role in plant growth. In this study, we investigated the response of Pistia stratiotes (P. stratiotes) to different concentrations of Zn and Cu, and the results showed that plant growth and photosynthesis were inhibited under both Zn (1, 2, 4, and 8 mg/L) and Cu (0.2, 0.4, 0.8, and 1 mg/L) stresses. The relative growth rates of P. stratiotes under 8 mg/L Zn or 1 mg/L Cu stress were 6.33% and 6.90%, which were much lower than those in the control group (10.86%). Meanwhile, Zn and Cu stress caused insignificant change in the relative water contents of plants. The decrease in phlorophyll fluorescence parameters and chlorophyll contents suggested the significant photoinhibition of Zn and Cu stress. Chemical analysis of plant root exudates showed that the root secretion species obtained by gas chromatography-mass spectrometry (GC-MS) mainly included amino acids, alkanes, aldehydes, ketones, phenols, and more. Compared with the control group, the influence of Zn or Cu on the reduction in relative amounts of exudates was greater than that on the increase. The results of this study provide important data for the utilization of P. stratiotes in heavy metal-polluted water environments.

Physiotyping of Plants and Modeling the Soil Plant Atmospheric Continuum (SPAC).

This chapter presents a holistic and quantitative approach to the carbon cycle in plant systems biology. It includes (rapid) phenotyping and monitoring of physiological key interactions of plants with its respective soil and atmospheric environment (soil plant atmospheric continuum-SPAC). The approach aims at qualifying and quantifying key components of this microhabitat as influenced by a single plant or a local group of plants in order to contribute to a flux-based modelling approach. The toolset consists of plant biometry, gas exchange, metabolomics, ionomics, root exudate characterization as well as soil biological and physical-chemical characterization. The results are presented as a basic interaction and input-output model aka conceptual system model employing H. T. Odum-style plots based on empirical data.

Root exudates and rhizosphere microbiota in responding to long-term continuous cropping of tobacco.

Soil sickness a severe problem in tobacco production, leading to soil-borne diseases and reduce in tobacco yield. This occurs as a result of the interaction between root exudates and rhizosphere microorganisms, which is however, little studied until now. By combining the field investigation and pot experiment, we found the output yield consistently decreased during the first 10 years of continuous cropping in a tobacco field, but increased at the 15th year (15Y). The root exudate and rhizosphere bacterial community was further analyzed to reveal the underlying mechanism of the suppressive soil formation. Root exudate of 15Y tobacco enriched in amino acids and derivatives, while depleted in the typical autotoxins including phenolic acids and alkaloids. This was correlated to the low microbial diversity in 15Y, but also the changes in community composition and topological properties of the co-occurrence network. Especially, the reduced autotoxins were associated with low Actinobacteria abundance, low network complexity and high network modularity, which significantly correlated with the recovered output yield in 15Y. This study revealed the coevolution of rhizosphere microbiota and root exudate as the soil domesticated by continuous cropping of tobacco, and indicated a potential role of the autotoxins and theirs effect on the microbial community in the formation of suppressive soil.

Neighbour-induced changes in root exudation patterns of buckwheat results in altered root architecture of redroot pigweed.

Roots are crucial in plant adaptation through the exudation of various compounds which are influenced and modified by environmental factors. Buckwheat root exudate and root system response to neighbouring plants (buckwheat or redroot pigweed) and how these exudates affect redroot pigweed was investigated. Characterising root exudates in plant-plant interactions presents challenges, therefore a split-root system which enabled the application of differential treatments to parts of a single root system and non-destructive sampling was developed. Non-targeted metabolome profiling revealed that neighbour presence and identity induces systemic changes. Buckwheat and redroot pigweed neighbour presence upregulated 64 and 46 metabolites, respectively, with an overlap of only 7 metabolites. Root morphology analysis showed that, while the presence of redroot pigweed decreased the number of root tips in buckwheat, buckwheat decreased total root length and volume, surface area, number of root tips, and forks of redroot pigweed. Treatment with exudates (from the roots of buckwheat and redroot pigweed closely interacting) on redroot pigweed decreased the total root length and number of forks of redroot pigweed seedlings when compared to controls. These findings provide understanding of how plants modify their root exudate composition in the presence of neighbours and how this impacts each other's root systems.

Harnessing root-soil-microbiota interactions for drought-resilient cereals.

Cereal plants form complex networks with their associated microbiome in the soil environment. A complex system including variations of numerous parameters of soil properties and host traits shapes the dynamics of cereal microbiota under drought. These multifaceted interactions can greatly affect carbon and nutrient cycling in soil and offer the potential to increase plant growth and fitness under drought conditions. Despite growing recognition of the importance of plant microbiota to agroecosystem functioning, harnessing the cereal root microbiota remains a significant challenge due to interacting and synergistic effects between root traits, soil properties, agricultural practices, and drought-related features. A better mechanistic understanding of root-soil-microbiota associations could lead to the development of novel strategies to improve cereal production under drought. In this review, we discuss the root-soil-microbiota interactions for improving the soil environment and host fitness under drought and suggest a roadmap for harnessing the benefits of these interactions for drought-resilient cereals. These methods include conservative trait-based approaches for the selection and breeding of plant genetic resources and manipulation of the soil environments.

Family ties: Root-root communication within Solanaceae.

Root-root communication effects on several physiological and metabolic aspects among Solanaceae relatives were studied. We examined cherry (C) and field (F) tomato (Solanum lycopersicum) and bell pepper (B) (Capsicum annuum), comprising three degrees of relatedness (DOR): high (H-DOR; CC, FF and BB), medium (M-DOR; CF) and low (L-DOR; CB and FB). Plants were grown in pairs of similar or different plants on a paper-based and non-destructive root growth system, namely, rhizoslides. Root growth, including the proliferation of fine roots, and respiration increased as the DOR decreased and were highest in paired L-DOR plants, as was shown for root respiration that increased by 63, 110 and 88 % for C, F, and B when grown with B, B and F, respectively. On the other hand, root exudates of L-DOR plants had significantly lower levels of total organic carbon and protein than those of H-DOR plants, indicating different root-root communication between individuals with different DOR. Our findings indicate, for the first time, that carbon allocation to root growth, exudation and respiration depends on the degree of genetic relatedness, and that the degree of relatedness between individual plants plays a key role in the root-root communication within Solanaceae.

Analysis of Ginkgo biloba Root Exudates and Inhibition of Soil Fungi by Flavonoids and Terpene Lactones.

Ginkgo biloba is abundant in secondary metabolites, including flavonoids and terpenoids. While the majority of research has focused on the role of these compounds in disease resistance, their specific contribution to pathogen defense has been rarely explored. In this study, we collected root exudates from hydroponically cultivated ginkgo seedlings and conducted a metabolomic analysis. We identified several primary metabolites mainly comprising amino acids and nucleotides, while secondary metabolites consisted of various compounds, including bioactive compounds such as flavonoids and terpenoids. Focusing on the secondary metabolites with relatively higher abundance in the exudates, we selected a mixture of flavonoids and terpenoids for in vitro inhibition experiments against two soil-borne fungal pathogens, Fusarium oxysporum f. sp. cucumerinum that causes cucumber wilt and Rhizoctonia solani AG-8 that causes wheat root rot. The results indicated that the growth rate of both fungus cells was significantly reduced with the increasing concentration of the flavonoid and terpenoid mixture extracted from ginkgo and was completely inhibited at a concentration of 5 mg/mL. Further experiments revealed that this mixture of flavonoids and terpenoids had a destructive effect on the cellular structure of both fungi, thereby reducing cell viability and achieving an antifungal effect. These findings provide a foundation for further research into the use of ginkgo extracts in biological control.

The dissipation of organophosphate esters mediated by ryegrass root exudate oxalic acid in soil: Analysis of enzymes activities, microorganism.

Complex rhizoremediation is the main mechanism of phytoremediation in organic-contaminated soil. Low molecular weight organic acids (LMWOAs) in root exudates have been shown to increase the bioavailability of contaminants and are essential for promoting the dissipation of contaminants. The effects of root exudates on the dissipation of organophosphate esters (OPEs) in soil are unclear. Consequently, we studied the combined effects of root exudates, soil enzymes and microorganisms on OPEs (tri (1-chloro-2-propyl) phosphate (TCPP) and triphenyl phosphate (TPP)) dissipation through pot experiments. Oxalic acid (OA) was confirmed to be the main component of LMWOAs in root exudates of ryegrass. The existence of OA increased the dissipation rate of OPEs by 6.04%-25.50%. Catalase and dehydrogenase activities were firstly activated and then inhibited in soil. While, urease activity was activated and alkaline phosphatase activity was inhibited during the exposure period. More bacteria enrichment (e.g., Sphingomonas, Pseudomonas, Flavisolibacter, Pontibacter, Methylophilus and Massilia) improved the biodegradation of OPEs. In addition, the transformation paths of OPEs hydrolysis and methylation under the action of root exudates were observed. This study provided theoretical insights into reducing the pollution risk of OPEs in the soil.

Evidence of nitrogen inputs affecting soil nitrogen purification by mediating root exudates of salt marsh plants.

Salt marsh has an important 'purification' role in coastal ecosystems by removing excess nitrogen that could otherwise harm aquatic life and reduce water quality. Recent studies suggest that salt marsh root exudates might be the 'control centre' for nitrogen transformation, but empirical evidence is lacking. Here we sought to estimate the direction and magnitude of nitrogen purification by salt marsh root exudates and gain a mechanistic understanding of the biogeochemical transformation pathway(s). To achieve this, we used a laboratory incubation to quantify both the root exudates and soil nitrogen purification rates, in addition to the enzyme activities and functional genes under Phragmites australis populations with different nitrogen forms addition (NO3-, NH4+ and urea). We found that NO3- and urea addition significantly stimulate P. australis root exudation of total acids, amino acids, total sugars and total organic carbon, while NH4+ addition only significantly increased total acids, amino acids and total phenol exudation. High total sugars, amino acids and total organic carbon concentrations enlarged nitrogen purification potential by stimulating the nitrogen purifying bacterial activities (including enzyme activities and related genes expression). Potential denitrification rates were not significantly elevated under NH4+ addition in comparison to NO3- and urea addition, which should be ascribed to total phenol self-toxicity and selective inhibition. Further, urea addition stimulated urease and protease activities with providing more NH4+ and NO2- substrates for elevated anaerobic ammonium oxidation rates among the nitrogen addition treatments. Overall, this study revealed that exogenous nitrogen could increase the nitrogen purification-associated bacterial activity through accelerating the root exudate release, which could stimulate the activity of nitrogen transformation, and then improve the nitrogen removal capacity in salt marsh.