Fungal Endophyte Phomopsis Research Articles

Root endophyte-enhanced peanut-rhizobia interaction is associated with regulation of root exudates

Root exudates play a crucial role in the symbiosis between leguminous plants and rhizobia. Our previous studies have shown that a fungal endophyte Phomopsis liquidambaris promotes peanut-rhizobia nodulation and nitrogen fixation, but the underlying mechanism are largely unknown. Here, we explore the role of peanut root exudates in Ph. liquidambaris-mediated nodulation enhancement. We first collected root exudates from Ph. liquidambaris-inoculated and un-inoculated peanuts and determined their effects on rhizobial growth, biofilm formation, chemotaxis, nodC gene expression, and peanut nodulation. Our results found a positive effect of Ph. liquidambaris-inoculated root exudates on these characteristics of rhizobia. Next, we compared the root exudates profile of Ph. liquidambaris-inoculated and un-inoculated plants and found that Ph. liquidambaris altered the concentrations of phenolic acids, flavonoids, organic acids and amino acids in root exudates. Furthermore, the rhizobial chemotaxis, growth and biofilm formation in response to the changed compounds at different concentrations showed that all of the test compounds induced rhizobial chemotactic behavior, and organic acids (citric acid and oxalic acid) and amino acid (glutamate, glycine and glutamine) at higher concentrations increased rhizobial growth and biofilm formation. Collectively, our results suggest that root exudates alterations contribute to Ph. liquidambaris-mediated peanut-rhizobia nodulation enhancement.

Carneic Acids from an Endophytic Phomopsis sp. as Dengue Virus Polymerase Inhibitors.

Thirteen carneic acids were isolated from the fungal endophyte Phomopsis sp. SNB-LAP1-7-32. Their structures were identified by mass spectrometry and extensive one- and two-dimensional NMR spectroscopy and through comparison with data reported in the literature. Compounds 1-13 were investigated for their antipolymerase activities against DENV polymerase and Zika NS5. Five of them exhibited significant inhibition of dengue polymerase with IC50 values in the 10 to 20 μM range without cytotoxicity. None inhibited Zika virus NS5 protein.

Phomopsis liquidambaris inoculation induces resistance in peanut to leaf spot and root rot

Soil-borne pathogens severely reduce the agricultural yield of peanuts. Accordingly, the application of biocontrol agents for plant disease management is a potentially powerful method for peanut cultivation. This study aimed to (1) evaluate the protective effect of the fungal endophyte Phomopsis liquidambaris for peanut plants against leaf spot disease caused by Colletotrichum gloeosporioides and against root rot disease caused by Fusarium oxysporum under outdoor conditions and (2) explore the potential biological control mechanisms of P. liquidambaris under greenhouse conditions. In our outdoor pot experiments, P. liquidambaris significantly reduced leaf spot and root rot diseases while increasing peanut yields. Under greenhouse conditions, P. liquidambaris–induced resistance against pathogens was associated with increased antioxidant enzyme activity, callose deposition, cell lignification, and phytoalexins accumulation. The demonstrated ability of P. liquidambaris to control disease makes it a strong biocontrol agent candidate.

Fungal endophyte Phomopsis liquidambari biodegrades soil resveratrol: a potential allelochemical in peanut monocropping systems

Most allelochemicals are secondary products released from root excretions or plant residues that accumulate in continuous cropping systems and cause severe decline in peanut yield. Resveratrol is a plant-derived stilbene that is released from peanut residues and accumulates in the soil; however, its allelopathic effects on peanut production are overlooked. Effective management solutions need to be developed to relieve allelopathy caused by soil resveratrol. Here, the biodegradation of resveratrol by the fungal endophyte Phomopsis liquidambari was investigated in a mineral salt medium and a soil trial. Resveratrol and its metabolites (produced by degradation by P. liquidambari) were detected by high-performance liquid chromatography-mass spectrometry (HPLC-MS). Resveratrol released from peanut residues reached a maximum concentration of 0.18 μg g-1 soil in litterbag experiments. Exogenous resveratrol inhibited peanut growth, nodule formation, and soil dehydrogenase activity, and reduced the soil microbial biomass carbon content and bacterial abundance, indicating an allelopathic role in peanut growth. More than 97% of the resveratrol was degraded within 72 and 168 h by P. liquidambari in pure culture and soil conditions, respectively. Resveratrol was first cleaved to 3,5-dihydroxybenzaldehyde and 4-hydroxybenzaldehyde, which were subsequently oxidized into 3,5-dihydroxybenzoic acid and 4-hydroxybenzoic acid, respectively. Fungal resveratrol cleavage oxygenase and the related gene expression were enhanced when P. liquidambari was induced by the resveratrol during the incubation. Our results indicate that the practical application of the fungal endophyte P. liquidambari has strong potential for biodegrading soil resveratrol, which can cause allelopathy in peanut continuous cropping systems. © 2019 Society of Chemical Industry.

Fungal Endophyte Phomopsis liquidambari Improves Iron and Molybdenum Nutrition Uptake of Peanut in Consecutive Monoculture Soil

Iron (Fe) and molybdenum (Mo) are essential trace elements for plants, especially for peanut. However, peanut planted in consecutive monoculture soil usually suffers from malnutrition. Thus, the objective of this study was to investigate the effects of the fungal endophyte Phomopsis liquidambari on Fe and Mo nutrition uptake of peanut in consecutive monoculture soil. Plant growth parameters, physiological parameters, the accumulation of Fe and Mo elements, gene expression related to absorption and transformation of Fe and Mo in planta, and the low-molecular-weight organic acids (LMWOAs) concentration in rhizosphere soil were analyzed by greenhouse pot experiments. The results showed that the application of P. liquidambari improved growth parameters, chlorophyll content, nitrate reductase (NR) activity, and Fe and Mo nutrition of peanut. Furthermore, LMWOAs in peanut rhizosphere soil and the expression levels of AhFRO1, AhIRT1, and AhMOT1 were significantly changed by the application of fungal endophyte. These findings indicated that the improvement of Fe and Mo nutrition in peanut was mostly derived from the promoting growth function of P. liquidambari and the regulation of peanut-P. liquidambari symbiosis with rhizosphere LMWOAs and Fe- and Mo-related gene expression in continuous monoculture soil.

Symbiotic fungal endophyte Phomopsis liquidambari-rice system promotes nitrogen transformation by influencing below-ground straw decomposition in paddy soil.

To explore if and how symbiotic Phomopsis liquidambari-rice system influences below-ground straw decomposition and then nitrogen(N) transformation in response to environmental N levels. Litter bag experiments were utilized to trace the decay process during rice growth phases (seedling (T1), tillering (T2), heading (T3) and maturing (T4) stage), with (E+) and without endophyte (E-), under low (LN), medium (MN) and high nitrogen (HN) supply. Litter, soil and plant samples were collected to evaluate the decay process, N transformations, plant quality and relative abundance of soil ammonia-oxidizing archaea (AOA), ammonia-oxidizing bacteria (AOB) and P. liquidambari. The results showed that straw decomposition increased by 19·76% (LN, T2 stage), 14·05% (MN, T3 stage) and 16·88% (MN, T4 stage) in E+ pots when compared with E- pots. Further analysis revealed that no significant endophyte×N interaction was found for straw decay rate and that the decay rate was reduced by a higher N supply (LN, 37·16±0·65%; MN, 32·27±1·72%; HN, 29·44±1·22%) at the T1 stage, whereas straw decay rate and N release increased by 9·38 and 11·16%, respectively, mainly by endophyte colonization at the T4 stage. The abundance of AOA and AOB were altered, corresponding with the decay rate. Soil mineral N, straw mineral N and plant quality were shown to increase in E+ pots, depending on environmental N conditions and growth phase. The yield increased by 2·98% for E+ plants under MN level. Symbiotic P. liquidambari-rice system promoted below-ground straw decomposition and N transformation, depending on environmental N levels and plant growth phase. This study provides evidence that fungal endophyte-plant systems are able to promote N transformation by increasing straw decomposition. A reasonable combination of N inputs could enhance its advantage in agriculture ecosystems.

Mutualistic fungus Phomopsis liquidambari increases root aerenchyma formation through auxin-mediated ethylene accumulation in rice (Oryza sativa L.)

The fungal endophyte Phomopsis liquidambari can improve nitrification rates and alter the abundance and composition of ammonia-oxidizers in the soil rhizosphere of rice. Aerenchyma is related to oxygen transport efficiency and contributes to the enhanced rhizospheric nitrification under flooding conditions. However, whether and how P. liquidambari affects aerenchyma formation is largely unknown. We therefore conducted pot and hydroponic experiments to investigate the changes of aerenchyma area, ethylene and indole-3-acetic acid (IAA) levels in rice with or without P. liquidambari infection. Our results showed that the larger aerenchyma area in rice roots with P. liquidambari inoculation was associated with markedly up-regulated expression of genes related to aerenchyma formation. Meanwhile, P. liquidambari inoculation substantially elevated root porosity (POR) and radial oxygen loss (ROL), leading to the enhancement of oxidation-reduction potential (ORP) under pot condition. Besides, P. liquidambari significantly increased IAA and ethylene levels in rice by stimulating the expression of genes involved in auxin and ethylene biosyntheses. Furthermore, auxin that partly acting upstream of ethylene signalling played an essential role in P. liquidambari-promoted aerenchyma formation. These results verified the direct contribution of P. liquidambari in promoting aerenchyma formation via the accumulation of IAA and ethylene in rice roots, which provides a constructive suggestion for improving hypoxia tolerance through plant-endophyte interactions.

Fungal endophyte Phomopsis liquidambari B3 enriches the diversity of nodular culturable endophytic bacteria associated with continuous cropping of peanut

ABSTRACTPreviously, the application of the fungal endophyte Phomopsis liquidambari B3 exhibited notable effect in alleviating soil obstacles caused by continuous cropping of peanut. It was supposed that P. liquidambari B3 could enhance the efficiency of nodulation and nitrogen fixation by enriching the diversity of nodular endophytic bacteria under continous monocropping system. To verify this hypothesis, a pot experiment was conducted under natural conditions. Four treatments were set as follow: discontinuous cropping soil (NS), continuous cropping soil (CK), continuous cropping soil inoculate with actived P. liquidambari B3 (CSB3), and continuous cropping soil inoculate with sterilized P. liquidambari B3 (CSIB3). As a result, a total of 120 isolates were obtained. Our study clearly declared that the addition of fungal endophyte effectively enriched the genetic diversity and the community composition in CSB3. In contrast, a low genetic diversity level and simplex community structure were exhibited in CK and CSIB3. Meanwhile, the positive percentage rate and corresponding community composition of the plant growth-promoting isolates in NS and CSB3 were larger and more abundant than the other. This is the first time to describe the effects of fungal endophytes on the diversity of the nodular culturable endophytic bacteria associated with continuous cropping of peanut.

Physiological mechanisms behind endophytic fungus Phomopsis liquidambari-mediated symbiosis enhancement of peanut in a monocropping system

Fungal endophyte Phomopsis liquidambari B3 effectively increases nodule number and productivity of peanut when grown in a monoculturing system, but the underlying mechanisms are not well understood. Plant physiological status is the key mechanism that determines the legume-rhizobium interaction under stressful conditions. Therefore, this research aimed to study the physiological mechanisms behind the P. liquidambari-mediated monoculturing peanut nodulation enhancement. Peanut-rhizobia symbiosis, plant defense enzyme activity in shoots and roots, photosynthetic activity and soluble sugar content in leaves, as well as the carbon metabolism-related enzyme activity and carbon metabolites content in nodules were measured after live P. liquidambari and P. liquidambari fragments treatment under continuous monoculturing condition. P. liquidambari instead of its fragments significantly enhanced nodule initiation, nodule development and nodule N2-fixation efficiency. Plants treated with live P. liquidambari showed higher leaf photosynthetic activity, soluble sugar accumulation, nodule carbohydrate catabolism activity and seed yield, whereas plant defense response was similar in endophyte and endophyte fragments treated plants. Our results demonstrated that the improved efficiency of symbiosis in peanut continuous monoculturing system, induced by P. liquidambari, was likely linked to the improved plant aboveground nutritional status rather than to the induction of plant defense.

Optimization of genome shuffling for high-yield production of the antitumor deacetylmycoepoxydiene in an endophytic fungus of mangrove plants.

As an accelerated evolutionary tool, genome shuffling is largely dependent on the high fusion frequency of different parental protoplasts. However, it was unclear how many types of parental protoplasts would afford the highest fusion frequency. Here, we applied the Monte Carlo method to simulate the simplified processes of protoplast fusion, to achieve maximal useful fusions in genome shuffling. The basic principle of this simulation is that valid fusions would take place when the minimum distance between two different types of parent protoplasts is smaller than that between two of the same types. Accordingly, simulations indicated that the highest fusion frequency would be achieved from eight to 12 different parental protoplasts. Based on the simulation results, eight parental protoplasts of the fungal endophyte Phomopsis sp. A123 were subjected to genome shuffling for yield improvement of deacetylmycoepoxydiene (DAM), an antitumor natural product with a novel chemical structure. After only two rounds of genome shuffling, four high-yield DAM-producing strains, namely G2-119, G2-448, G2-866, and G2-919, were obtained with the aid of activity screening and HPLC analysis. The results showed that the DAM yield in these four strains were 243-, 241-, 225-, and 275-fold, respectively, higher than that of the starting strain A123. This is the first time Monte Carlo simulation is introduced into the field of cell fusion and is also the first report on the optimization of genome shuffling focusing on the number of parental types in protoplast fusions.

Enhanced nodulation of peanut when co-inoculated with fungal endophyte Phomopsis liquidambari and bradyrhizobium

In peanut continuous cropping soil, the application of fungal endophyte Phomopsis liquidambari B3 showed peanut pod yield promotion and root nodule number increase. P. liquidambari improved soil environment by degrading allelochemicals and thus promoted peanut pod yield. Furthermore, peanut yield promotion is in part due to the root nodule increase since nodular nitrogen fixation provides the largest source of nitrogen for peanut. However, it is unknown whether this nodule number increase is induced by fungal endophyte. We therefore conducted several pot experiments using vermiculite to investigate the effects of P. liquidambari on peanut-bradyrhizobium nodulation. Our results showed that P. liquidambari co-inoculated with bradyrhizobium increased root nodule number and shoot accumulated nitrogen by 28.25% and 29.71%, respectively. Nodulation dynamics analysis showed that P. liquidambari accelerated nodule initiation and subsequent nodule development. Meanwhile, P. liquidambari was able to colonize the peanut root as an endophyte. The dynamics of P. liquidambari and bradyrhizobial root colonization analysis showed that P. liquidambari inoculation significantly increased the rate of bradyrhizobial colonization. Furthermore, P. liquidambari inoculation significantly increased flavonoids synthesis-related enzymes activities, two common types of flavonoid (luteolin and quercetin-peanut rhizobial nod gene inducer) secretion and lateral root (peanut rhizobial infection site) formation, indicating that P. liquidambari altered the peanut nodulation-related physiological and metabolic activities. These obtained results confirmed the direct contribution of P. liquidambari in enhancing peanut-bradyrhizobium interaction, nodulation and yield.

Fungal endophyte Phomopsis liquidambari affects nitrogen transformation processes and related microorganisms in the rice rhizosphere.

The endophytic fungus Phomopsis liquidambari performs an important ecosystem service by assisting its host with acquiring soil nitrogen (N), but little is known regarding how this fungus influences soil N nutrient properties and microbial communities. In this study, we investigated the impact of P. liquidambari on N dynamics, the abundance and composition of N cycling genes in rhizosphere soil treated with three levels of N (urea). Ammonia-oxidizing archaea (AOA), ammonia-oxidizing bacteria (AOB) and diazotrophs were assayed using quantitative real-time polymerase chain reaction and denaturing gradient gel electrophoresis at four rice growing stages (S0: before planting, S1: tillering stage, S2: grain filling stage, and S3: ripening stage). A significant increase in the available nitrate and ammonium contents was found in the rhizosphere soil of endophyte-infected rice under low N conditions. Moreover, P. liquidambari significantly increased the potential nitrification rates, affected the abundance and community structure of AOA, AOB, and diazotrophs under low N conditions in the S1 and S2 stages. The root exudates were determined due to their important role in rhizosphere interactions. P. liquidambari colonization altered the exudation of organic compounds by rice roots and P. liquidambari increased the concentration of soluble saccharides, total free amino acids and organic acids in root exudates. Plant-soil feedback mechanisms may be mediated by the rice-endophyte interaction, especially in nutrient-limited soil.

Improvement of nitrogen accumulation and metabolism in rice (Oryza sativa L.) by the endophyte Phomopsis liquidambari

The fungal endophyte Phomopsis liquidambari can enhance nitrogen (N) uptake and metabolism of rice plants under hydroponic conditions. To investigate the effects of P. liquidambari on N accumulation and metabolism in rice (Oryza sativa L.) under field conditions during the entire growing season (S1, the seedling stage; S2, the tillering stage; S3, the heading stage; S4, the ripening stage), we utilized pot experiments to examine metabolic and physiological levels in both shoot and root tissues of rice, with endophyte (E+) and without endophyte (E−), in response to three different N levels. We found that under low-N treatment, P. liquidambari symbiosis increased the rice yield and N use efficiency by 12% and by 11.59%, respectively; that the total N contents in E+ rice plants at the four growth stages were separately increased by 29.05%, 14.65%, 21.06% and 18.38%, respectively; and that the activities of nitrate reductase and glutamine synthetase in E+ rice roots and shoots were significantly increased by fungal infection during the S1 to S3 stages. Moreover, P. liquidambari significantly increased the free NH4+, NO3-, amino acid and soluble protein contents in infected rice tissues under low-N treatment during the S1 to S3 stages. The obtained results offer novel data concerning the systemic changes induced by P. liquidambari in rice during the entire growth period and confirm the hypothesis that the rice–P. liquidambari interaction improved the N accumulation and metabolism of rice plants, consequently increasing rice N utilization in nutrient-limited soil.

Effects of the fungal endophyte Phomopsis liquidambari on nitrogen uptake and metabolism in rice

Phomopsis liquidambari can establish a mutualistic symbiotic relationship with rice. It promotes the growth and yield of the host plant and reduces the amount of nitrogen (N) fertilizer required for plant growth. However, the mechanisms responsible for the effects of the fungal endophyte on N use in rice are largely unknown. We conducted a hydroponic experiment to investigate the effects of P. liquidambari on N uptake and N metabolism in rice plants. Rice plants were cultivated in the presence or absence of P. liquidambari under three N levels. Under the low-N treatment, fungal infection significantly increased the biomass, and the total N, soluble protein, free amino acid, free NH4 +, and chlorophyll contents of rice roots and shoots. The activities of nitrate reductase and glutamine synthetase were increased in infected rice plants. Some genes related to N uptake (OsAMT1;1, OsAMT1;3, OsAMT2;2, OsAMT3;2, OsAMT3;3, OsNRT2;1) and N metabolism (OsNR1, OsGS1, OsGS2, OsNADH-GOGAT) were also up-regulated in infected plants under the low-N treatment. However, these effects gradually weakened as the N level increased. The colonization rate of the endophyte substantially decreased with increasing N levels. Taken together, these results suggest that low-N fertilization induces a physiological state in rice that is favorable for the P. liquidambari symbiosis. The greater extent of P. liquidambari colonization under low-N conditions stimulated the expression of several genes involved in N uptake and N metabolism in rice, thereby enhancing N utilization. These results have implications for enhancing plant growth in low-input systems at nutrient-poor sites.