Beneficial Plant-microbe Interactions Research Articles

Ecosystem Fabrication (EcoFAB) Protocols for The Construction of Laboratory Ecosystems Designed to Study Plant-microbe Interactions

Beneficial plant-microbe interactions offer a sustainable biological solution with the potential to boost low-input food and bioenergy production. A better mechanistic understanding of these complex plant-microbe interactions will be crucial to improving plant production as well as performing basic ecological studies investigating plant-soil-microbe interactions. Here, a detailed description for ecosystem fabrication is presented, using widely available 3D printing technologies, to create controlled laboratory habitats (EcoFABs) for mechanistic studies of plant-microbe interactions within specific environmental conditions. Two sizes of EcoFABs are described that are suited for the investigation of microbial interactions with various plant species, including Arabidopsis thaliana, Brachypodium distachyon, and Panicum virgatum. These flow-through devices allow for controlled manipulation and sampling of root microbiomes, root chemistry as well as imaging of root morphology and microbial localization. This protocol includes the details for maintaining sterile conditions inside EcoFABs and mounting independent LED light systems onto EcoFABs. Detailed methods for addition of different forms of media, including soils, sand, and liquid growth media coupled to the characterization of these systems using imaging and metabolomics are described. Together, these systems enable dynamic and detailed investigation of plant and plant-microbial consortia including the manipulation of microbiome composition (including mutants), the monitoring of plant growth, root morphology, exudate composition, and microbial localization under controlled environmental conditions. We anticipate that these detailed protocols will serve as an important starting point for other researchers, ideally helping create standardized experimental systems for investigating plant-microbe interactions.

Tiny Microbes, Big Yields: enhancing food crop production with biological solutions.

SummaryPlant‐associated microbiomes have tremendous potential to improve plant resilience and yields in farming systems. There is increasing evidence that biological technologies that use microbes or their metabolites can enhance nutrient uptake and yield, control pests and mitigate plant stress responses. However, to fully realize the potential of microbial technology, their efficacy and consistency under the broad range of real‐world conditions need to be improved. While the optimization of microbial biofertilizers and biopesticides is advancing rapidly to enable use in various soils, crop varieties and environments, crop breeding programmes have yet to incorporate the selection of beneficial plant–microbe interactions to breed ‘microbe‐optimized plants’. Emerging efforts exploring microbiome engineering could lead to microbial consortia that are better suited to support plants. The combination of all three approaches could be integrated to achieve maximum benefits and significantly improved crop yields to address food security.

Third-party mutualists have contrasting effects on host invasion under the enemy-release and biotic-resistance hypotheses

Plants engage in complex multipartite interactions with mutualists and antagonists, but these interactions are rarely included in studies that explore plant invasiveness. When considered in isolation, we know that beneficial microbes can enhance an exotic plant’s invasive ability and that herbivorous insects often decrease an exotic plant’s likeliness of success. However, the effect of these partners on plant fitness has not been well characterized when all three species coevolve. We use computational evolutionary modeling of a trait-based system to test how microbes and herbivores simultaneously coevolving with an invading plant affect the invaders’ probability of becoming established. Specifically, we designed a model that explores how a beneficial microbe would influence the outcome of an interaction between a plant and herbivore. To model novel interactions, we included a phenotypic trait shared by each species. Making this trait continuous and selectable allows us to explore how trait similarities between coevolving plants, herbivores and microbes affect fitness. Using this model, we answer the following questions: (1) Can a beneficial plant-microbe interaction influence the evolutionary outcome of antagonistic interactions between plants and herbivores? (2) How does the initial trait similarity between interacting organisms affect the likelihood of plant survival in novel locations? (3) Does the effect of tripartite interactions on the invasion success of a plant depend on whether organisms interact through trait similarity [Enemy Release Hypothesis (ERH)] or dissimilarity (Biotic Resistance Hypothesis)? We found that it was much more difficult for plants to invade under the ERH but that beneficial microbes increase the probability of plant survival in a novel range under both hypotheses. To our knowledge, this model is the first to use tripartite interactions to explore novel species introductions. It represents a step towards gaining a better understanding of the factors influencing establishment of exotic species to prevent future invasions.

Iron and Immunity.

Iron is an essential nutrient for most life on Earth because it functions as a crucial redox catalyst in many cellular processes. However, when present in excess iron can lead to the formation of harmful hydroxyl radicals. Hence, the cellular iron balance must be tightly controlled. Perturbation of iron homeostasis is a major strategy in host-pathogen interactions. Plants use iron-withholding strategies to reduce pathogen virulence or to locally increase iron levels to activate a toxic oxidative burst. Some plant pathogens counteract such defenses by secreting iron-scavenging siderophores that promote iron uptake and alleviate iron-regulated host immune responses. Mutualistic root microbiota can also influence plant disease via iron. They compete for iron with soil-borne pathogens or induce a systemic resistance that shares early signaling components with the root iron-uptake machinery. This review describes the progress in our understanding of the role of iron homeostasis in both pathogenic and beneficial plant-microbe interactions.

Perspectives and Challenges of Microbial Application for Crop Improvement.

Global population increases and climate change pose a challenge to worldwide crop production. There is a need to intensify agricultural production in a sustainable manner and to find solutions to combat abiotic stress, pathogens, and pests. Plants are associated with complex microbiomes, which have an ability to promote plant growth and stress tolerance, support plant nutrition, and antagonize plant pathogens. The integration of beneficial plant-microbe and microbiome interactions may represent a promising sustainable solution to improve agricultural production. The widespread commercial use of the plant beneficial microorganisms will require a number of issues addressed. Systems approach using microscale information technology for microbiome metabolic reconstruction has potential to advance the microbial reproducible application under natural conditions.

Down-Regulation of KORRIGAN-Like Endo-β-1,4-Glucanase Genes Impacts Carbon Partitioning, Mycorrhizal Colonization and Biomass Production in Populus.

A greater understanding of the genetic regulation of plant cell wall remodeling and the impact of modified cell walls on plant performance is important for the development of sustainable biofuel crops. Here, we studied the impact of down-regulating KORRIGAN-like cell wall biosynthesis genes, belonging to the endo-β-1,4-glucanase gene family, on Populus growth, metabolism and the ability to interact with symbiotic microbes. The reductions in cellulose content and lignin syringyl-to-guaiacyl unit ratio, and increase in cellulose crystallinity of cell walls of PdKOR RNAi plants corroborated the functional role of PdKOR in cell wall biosynthesis. Altered metabolism and reduced growth characteristics of RNAi plants revealed new implications on carbon allocation and partitioning. The distinctive metabolome phenotype comprised of a higher phenolic and salicylic acid content, and reduced lignin, shikimic acid and maleic acid content relative to control. Plant sustainability implications of modified cell walls on beneficial plant-microbe interactions were explored via co-culture with an ectomycorrhizal fungus, Laccaria bicolor. A significant increase in the mycorrhization rate was observed in transgenic plants, leading to measurable beneficial growth effects. These findings present new evidence for functional interconnectedness of cellulose biosynthesis pathway, metabolism and mycorrhizal association in plants, and further emphasize the consideration of the sustainability implications of plant trait improvement efforts.

A perspective on inter-kingdom signaling in plant-beneficial microbe interactions.

Recent work has shown that the rhizospheric and phyllospheric microbiomes of plants are composed of highly diverse microbial species. Though the information pertaining to the diversity of the aboveground and belowground microbes associated with plants is known, an understanding of the mechanisms by which these diverse microbes function is still in its infancy. Plants are sessile organisms, that depend upon chemical signals to interact with the microbiota. Of late, the studies related to the impact of microbes on plants have gained much traction in the research literature, supporting diverse functional roles of microbes on plant health. However, how these microbes interact as a community to confer beneficial traits to plants is still poorly understood. Recent advances in the use of "biologicals" as bio-fertilizers and biocontrol agents for sustainable agricultural practices is promising, and a fundamental understanding of how microbes in community work on plants could help this approach be more successful. This review attempts to highlight the importance of different signaling events that mediate a beneficial plant microbe interaction. Fundamental research is needed to understand how plants react to different benign microbes and how these microbes are interacting with each other. This review highlights the importance of chemical signaling, and biochemical and genetic events which determine the efficacy of benign microbes to promote the development of beneficial traits in plants.

The core microbiome bonds the Alpine bog vegetation to a transkingdom metacommunity.

Bog ecosystems fulfil important functions in Earth's carbon and water turnover. While plant communities and their keystone species Sphagnum have been well studied, less is known about the microbial communities associated with them. To study our hypothesis that bog plants share an essential core of their microbiome despite their different phylogenetic origins, we analysed four plant community plots with 24 bryophytes, vascular plants and lichen species in two Alpine bogs in Austria by 16S rDNA amplicon sequencing followed by bioinformatic analyses. The overall bog microbiome was classified into 32 microbial phyla, while Proteobacteria (30.8%), Verrucomicrobia (20.3%) and Planctomycetes (15.1%) belonged to the most abundant groups. Interestingly, the archaeal phylum Euryarcheota represented 7.2% of total microbial abundance. However, a high portion of micro-organisms remained unassigned at phylum and class level, respectively. The core microbiome of the bog vegetation contained 177 operational taxonomic units (OTUs) (150526seq.) and contributed to 49.5% of the total microbial abundance. Only a minor portion of associated core micro-organisms was host specific for examined plant groups (5.9-11.6%). Using our new approach to analyse plant-microbial communities in an integral framework of ecosystem, vegetation and microbiome, we demonstrated that bog vegetation harboured a core microbiome that is shared between plants and lichens over the whole ecosystem and formed a transkingdom metacommunity. All micro- and macro-organisms are connected to keystone Sphagnum mosses via set of microbial species, for example Burkholderia bryophila which was found associated with a wide spectrum of host plants and is known for a beneficial plant-microbe interaction.

Biological management of Sclerotinia sclerotiorum in pea using plant growth promoting microbial consortium.

The beneficial plant-microbe interactions play crucial roles in protection against large number of plant pathogens causing disease. The present study aims to investigate the growth promoting traits induced by beneficial microbes namely Pseudomonas aeruginosa PJHU15, Trichoderma harzianum TNHU27, and Bacillus subtilis BHHU100 treated singly and in combinations under greenhouse and field conditions to control Sclerotinia sclerotiorum. Plants treated with three microbe consortium enhanced plant growth maximally both in the presence and absence of the pathogen. Increase in plant length, total biomass, number of leaves, nodules and secondary roots, total chlorophyll and carotenoid content, and yield were recorded in plants treated with microbial consortia. Also, a decrease in plant mortality was observed in plants treated with microbial consortia in comparison to untreated control plants challenged with S. sclerotiorum. Furthermore, the decrease in disease of all the treatments can be associated with differential improvement of growth induced in pea.

Paenibacillus yonginensis DCY84T induces changes in Arabidopsis thaliana gene expression against aluminum, drought, and salt stress

Current agricultural production methods, for example the improper use of chemical fertilizers and pesticides, create many health and environmental problems. Use of plant growth-promoting bacteria (PGPB) for agricultural benefits is increasing worldwide and also appears to be a trend for the future. There is possibility to develop microbial inoculants for use in agricultural biotechnology, based on these beneficial plant–microbe interactions. For this study, ten bacterial strains were isolated from Yongin forest soil for which in vitro plant-growth promoting trait screenings, such as indole acetic acid (IAA) production, a phosphate solubilization test, and a siderophore production test were used to select two PGPB candidates. Arabidopsis thaliana plants were inoculated with Paenibacillus yonginensis DCY84T and Micrococcus yunnanensis PGPB7. Salt stress, drought stress and heavy metal (aluminum) stress challenges indicated that P. yonginensis DCY84T-inoculated plants were more resistant than control plants. AtRSA1, AtVQ9 and AtWRKY8 were used as the salinity responsive genes. The AtERD15, AtRAB18, and AtLT178 were selected to check A. thaliana responses to drought stress. Aluminum stress response was checked using AtAIP, AtALS3 and AtALMT1. The qRT-PCR results indicated that P. yonginensis DCY84T can promote plant tolerance against salt, drought, and aluminum stress. P. yonginensis DCY84T also showed positive results during in vitro compatibility testing and virulence assay against X. oryzae pv. oryzae Philippine race 6 (PXO99). Better germination rates and growth parameters were also recorded for the P. yonginensis DCY84T Chuchung cultivar rice seed which was grown on coastal soil collected from Suncheon. Based on these results, P. yonginensis DCY84T can be used as a promising PGPB isolate for crop improvement.

21st century agriculture: integration of plant microbiomes for improved crop production and food security.

21st century agriculture: integration of plant microbiomes for improved crop production and food security.

Understanding and engineering beneficial plant-microbe interactions: plant growth promotion in energy crops.

Plant production systems globally must be optimized to produce stable high yields from limited land under changing and variable climates. Demands for food, animal feed, and feedstocks for bioenergy and biorefining applications, are increasing with population growth, urbanization and affluence. Low-input, sustainable, alternatives to petrochemical-derived fertilizers and pesticides are required to reduce input costs and maintain or increase yields, with potential biological solutions having an important role to play. In contrast to crops that have been bred for food, many bioenergy crops are largely undomesticated, and so there is an opportunity to harness beneficial plant–microbe relationships which may have been inadvertently lost through intensive crop breeding. Plant–microbe interactions span a wide range of relationships in which one or both of the organisms may have a beneficial, neutral or negative effect on the other partner. A relatively small number of beneficial plant–microbe interactions are well understood and already exploited; however, others remain understudied and represent an untapped reservoir for optimizing plant production. There may be near-term applications for bacterial strains as microbial biopesticides and biofertilizers to increase biomass yield from energy crops grown on land unsuitable for food production. Longer term aims involve the design of synthetic genetic circuits within and between the host and microbes to optimize plant production. A highly exciting prospect is that endosymbionts comprise a unique resource of reduced complexity microbial genomes with adaptive traits of great interest for a wide variety of applications.

Transcriptional regulation of Arabidopsis thaliana WRKY genes under interaction with beneficial fungus Trichoderma atroviride

Plants are associated with a wide range of microorganisms, and these interactions induce changes in both the plant and microorganism. Transcription factors play an important role in the regulation of large numbers of genes associated to plant–microbe response. WRKY transcription factors have been involved in the responses to plant–pathogen interactions, but little is known about WRKY transcription factors in beneficial plant–microbe interactions. In this study, the expression patterns of Arabidopsis thalianaWRKY genes were evaluated during the interaction with the beneficial fungus Trichoderma atroviride. Eight WRKY genes, AtWRKY8, AtWRKY33, AtWRKY38, AtWRKY42, AtWRKY54, AtWRKY57, AtWRKY60 and AtWRKY70, were analyzed by quantitative RT-PCR. These WRKY genes were found differentially expressed in a time-dependent manner during T. atroviride interaction. Our data suggest that T. atroviride induces the expression of positive regulators in jasmonic acid-mediated pathway such as AtWRKY8, AtWRKY33, AtWRKY38, AtWRKY42 and AtWRKY60, while salicylic acid pathway regulated by AtWRKY70 and AtWRKY54, could be activated at later stages of the interaction, when the fungus is fully established in the plant roots. In addition, Trichoderma treatment regulates the expression of WRKY genes such as AtWRKY57, AtWRKY60 and AtWRKY33 related to response to abiotic stresses. In this sense, WRKY transcription factors regulation suggests a complex signaling network in this beneficial plant–microbe interaction.

Effects of bacterial inoculants on the indigenous microbiome and secondary metabolites of chamomile plants

Plant-associated bacteria fulfill important functions for plant growth and health. However, our knowledge about the impact of bacterial treatments on the host's microbiome and physiology is limited. The present study was conducted to assess the impact of bacterial inoculants on the microbiome of chamomile plants Chamomilla recutita (L.) Rauschert grown in a field under organic management in Egypt. Chamomile seedlings were inoculated with three indigenous Gram-positive strains (Streptomyces subrutilus Wbn2-11, Bacillus subtilis Co1-6, Paenibacillus polymyxa Mc5Re-14) from Egypt and three European Gram-negative strains (Pseudomonas fluorescens L13-6-12, Stenotrophomonas rhizophila P69, Serratia plymuthica 3Re4-18) already known for their beneficial plant-microbe interaction. Molecular fingerprints of 16S rRNA gene as well as real-time PCR analyses did not show statistically significant differences for all applied bacterial antagonists compared to the control. In contrast, a pyrosequencing analysis of the 16S rRNA gene libraries revealed significant differences in the community structure of bacteria between the treatments. These differences could be clearly shown by a shift within the community structure and corresponding beta-diversity indices. Moreover, B. subtilis Co1-6 and P. polymyxa Mc5Re-14 showed an enhancement of the bioactive secondary metabolite apigenin-7-O-glucoside. This indicates a possible new function of bacterial inoculants: to interact with the plant microbiome as well as to influence the plant metabolome.

Biocontrol Agent Talaromyces flavus Stimulates the Growth of Cotton and Potato

Beneficial plant-microbe interactions in the rhizosphere are primary determinants of plant health and soil fertility. Some antagonistic fungi have shown great effects toward the growth of plant crops. In this study, two major crops, cotton and potato, were selected to evaluate their growth promotion by the antagonistic fungus Talaromyces flavus. For each plant, five T. flavus isolates were selected from our fungal collection which had shown the highest antagonistic activities against the causal agent of wilt diseases on these plants. In the next step, for every crop, five isolates were used under greenhouse conditions. For evaluation of the plant growth promotion ability of T. flavus isolates, a split-plot trial was arranged in a randomized complete block design with four replications. The main factor was the method of application of T. flavus as a soil treatment, a seed treatment, and a combination of both methods. The subfactor was the use of different fungal isolates. Measured parameters were root length, crown length, plant height, plant fresh weight, and plant dry weight. Results showed that the maximum increase in the above parameters was mediated by the seed treatment method. The most effective isolate for cotton plants was TF-Co-M-23, which increased root length, plant height, plant fresh weight, and plant dry weight by 1.80-, 2.26-, 1.23-, and 1.19-fold, respectively. There were no significant differences among the various treatments affected by T. flavus in terms of crown length. The most effective isolate for potato plants was TF-Po-V-50, which increased root length, crown length, plant height, and plant dry weight by 1.71-, 1.09-, 1.45-, and 3.75-fold, respectively. The overall results of this study suggest that it may be possible to promote cotton and potato growth characteristics by using the antagonistic fungus T. flavus.

Role of the RNA-binding protein Hfq in Serratia plymuthica

The RNA-binding protein Hfq has been well studied as a global post-transcriptional regulator which controls diverse cellular processes in bacteria. However, the function in the genus of Serratia has remained unexplored. Here we show that beyond mutation in Hfq resulting in their growth defects, Hfq has global effects on a variety of biocontrol-related phenotypes in the endophytic strain G3 of Serratia plymuthica, including antifungal activity, production of exoenzymes, as well as motility and biofilm formation. Especially for the first time, Hfq is observed to control biosynthesis of auxin indole-3-acetic acid (IAA) and antibiotic pyrrolnitrin (PRN), which are key determinants responsible for plant growth promotion and suppression of phytopathogens, respectively by G3. Additionally, Hfq is also required for the production of RpoS, a major stress sigma factor in G3. In contrast to E. coli, translation of hfq in G3 is positively autoregulated. Further investigation of the molecular mechanisms involved in regulation of IAA and pyrrolnitrin production by Hfq and its role in the regulatory networks of G3 will help to optimize the beneficial plant-microbe interactions.

Genetic Diversity of Cultivable Plant Growth-Promoting Rhizobacteria in Korea

To elucidate the biodiversity of plant growth-promoting rhizobacteria (PGPR) in Korea, 7,638 bacteria isolated from the rhizosphere of plant species growing in many different regions were screened. A large number of PGPR were identified by testing the ability of each isolate to promote the growth of cucumber seedlings. After redundant rhizobacteria were removed via amplified rDNA restriction analysis, 90 strains were finally selected as PGPR. On the basis of 16S ribosomal RNA sequences, 68 Gram-positive (76%) and 22 Gram-negative (24%) isolates were assigned to 21 genera and 47 species. Of these genera, Bacillus (32 species) made up the largest complement, followed by Paenibacillus (19) and Pseudomonas (11). Phylogenetic analysis showed that most of the Grampositive PGPR fell into two categories: low- and high- G+C (Actinobacteria) strains. The Gram-negative PGPR were distributed in three categories: alpha-proteobacteria, beta- proteobacteria, and gamma-proteobacteria. To our knowledge, this is the largest screening study designed to isolate diverse PGPR. The enlarged understanding of PGPR genetic diversity provided herein will expand the knowledge base regarding beneficial plant-microbe interactions. The outcome of this research may have a practical effect on crop production methodologies.

Plant defense: warding off attack by pathogens, herbivores, and parasitic plants

Plant defense: warding off attack by pathogens, herbivores, and parasitic plants

Elicitation of foliar resistance mechanisms transiently impairs root association with arbuscular mycorrhizal fungi

Summary 1. Plants possess numerous mechanisms to control infections by deleterious organisms. Unspecific resistance mechanisms may, however, also exert ecological costs when they have a negative effect on beneficial plant–microbe interactions. Such negative effects may even cross the border between a plant’s aerial parts and its roots and then affect very central functions such as nutrient uptake and root resistance to micro‐organisms. Whereas an impaired nodulation indeed appears common after resistance expression in the leaves, contradictory results have been published for the case of arbuscular mycorrhizal (AM) fungi. 2. We analysed the effect of induction of resistance mechanisms in foliar tissues on AM colonization in soybean plants, using acibenzolar‐S‐methyl (ASM) as chemical elicitor. By determining different physiological and biochemical parameters, we assessed whether the effects are related to the activation of the plant defence mechanisms or rather to the re‐allocation of primary metabolites. 3. Colonization with AM fungi transiently decreased after pathogen resistance mechanisms were elicited in the aerial parts of the plant. The induction with ASM led to a significant, yet moderate, defence response in the roots, which was modulated in mycorrhizal plants. No allocation or fitness costs associated with the induction of resistance were detected in this study. 4. Synthesis. Our study confirms a transient negative impact of the elicitation of foliar defences on root–AM interactions. The results show that induced resistance to foliar pathogens can (i) move from the above‐ground to the below‐ground compartment and (ii) affect mutualistic micro‐organisms as well as plant pathogens. We also conclude that (iii) the negative effect is likely linked to changes in the defence status of the plant rather than to changes in resource allocation patterns and (iv) the AM association can modulate the activation of the plant defence mechanisms and overcome such effects.

Transcription profiling of the isoflavone phenylpropanoid pathway in soybean in response to Bradyrhizobium japonicum inoculation

Isoflavones are legume-specific secondary metabolites that function as defence compounds, signal molecules and regulators of gene expression during both pathogen attack and beneficial plant-microbe interactions. They are synthesised by a branch of the core phenylpropanoid pathway, using several isoenzymes within each enzymatic step. Gene-specific quantitative real-time reverse transcriptase PCR (qRT-PCR) was used to quantify expression of isoflavone synthesis genes in soybean (Glycine max L). Genes encoding chalcone synthase 7 (CHS7), chalcone synthase 8 (CHS8) and isoflavone synthase 1 (IFS1) displayed high basal expression levels in roots compared with hypocotyls, suggesting they could be the gene family members encoding the isoenzyme that contributes the most to the principal substrate flux towards specific isoflavone synthesis in roots. The genes encoding phenylalanine ammonia lyase 1 (PAL1) and IFS1 showed induction in root tissue after inoculation with Bradyrhizobium japonicum (Kirchner) Jordan, suggesting a control point. The absence of a functional nodulation regulator, GmNARK (G. max nodulation autoregulation receptor kinase), in the soybean mutant nts1007 resulted in significantly increased basal expression of PAL1 compared with levels induced by B. japonicum, suggesting that GmNARK is a negative regulator for isoflavone phenylpropanoid pathway genes during nodulation and that distinct genes, as opposed to the complete pathway, are coordinately regulated by the nodulation status of the mutant.