Beneficial Plant-microbe Interactions Research Articles

Culture-based Methods for Studying the Bacterial Root Microbiome of Wheat.

Beneficial plant-microbe interactions are important and desirable for sustainable intensification of agriculture. Here, we describe methods to isolate microbes from the roots of field-grown wheat plants. This includes the rhizosphere and rhizoplane soil, as well as the root endosphere. We also describe a method to test for endosphere competence of putative endophytes.

Heritable Variation in Pea for Resistance Against a Root Rot Complex and Its Characterization by Amplicon Sequencing.

Soil-borne pathogens cause severe root rot of pea (Pisum sativum L.) and are a major constraint to pea cultivation worldwide. Resistance against individual pathogen species is often ineffective in the field where multiple pathogens form a pea root rot complex (PRRC) and conjointly infect pea plants. On the other hand, various beneficial plant-microbe interactions are known that offer opportunities to strengthen plant health. To account for the whole rhizosphere microbiome in the assessment of root rot resistance in pea, an infested soil-based resistance screening assay was established. The infested soil originated from a field that showed severe pea root rot in the past. Initially, amplicon sequencing was employed to characterize the fungal microbiome of diseased pea roots grown in the infested soil. The amplicon sequencing evidenced a diverse fungal community in the roots including pea pathogens Fusarium oxysporum, F. solani, Didymella sp., and Rhizoctonia solani and antagonists such as Clonostachys rosea and several mycorrhizal species. The screening system allowed for a reproducible assessment of disease parameters among 261 pea cultivars, breeding lines, and landraces grown for 21 days under controlled conditions. A sterile soil control treatment was used to calculate relative shoot and root biomass in order to compare growth performance of pea lines with highly different growth morphologies. Broad sense heritability was calculated from linear mixed model estimated variance components for all traits. Emergence on the infested soil showed high (H2 = 0.89), root rot index (H2 = 0.43), and relative shoot dry weight (H2 = 0.51) medium heritability. The resistance screening allowed for a reproducible distinction between PRRC susceptible and resistant pea lines. The combined assessment of root rot index and relative shoot dry weight allowed to identify resistant (low root rot index) and tolerant pea lines (low relative shoot dry weight at moderate to high root rot index). We conclude that relative shoot dry weight is a valuable trait to select disease tolerant pea lines. Subsequently, the resistance ranking was verified in an on-farm experiment with a subset of pea lines. We found a significant correlation (rs = 0.73, p = 0.03) between the controlled conditions and the resistance ranking in a field with high PRRC infestation. The screening system allows to predict PRRC resistance for a given field site and offers a tool for selection at the seedling stage in breeding nurseries. Using the complexity of the infested field soil, the screening system provides opportunities to study plant resistance in the light of diverse plant-microbe interactions occurring in the rhizosphere.

Soil origin and plant genotype structure distinct microbiome compartments in the model legume Medicago truncatula

BackgroundUnderstanding the genetic and environmental factors that structure plant microbiomes is necessary for leveraging these interactions to address critical needs in agriculture, conservation, and sustainability. Legumes, which form root nodule symbioses with nitrogen-fixing rhizobia, have served as model plants for understanding the genetics and evolution of beneficial plant-microbe interactions for decades, and thus have added value as models of plant-microbiome interactions. Here we use a common garden experiment with 16S rRNA gene amplicon and shotgun metagenomic sequencing to study the drivers of microbiome diversity and composition in three genotypes of the model legume Medicago truncatula grown in two native soil communities.ResultsBacterial diversity decreased between external (rhizosphere) and internal plant compartments (root endosphere, nodule endosphere, and leaf endosphere). Community composition was shaped by strong compartment × soil origin and compartment × plant genotype interactions, driven by significant soil origin effects in the rhizosphere and significant plant genotype effects in the root endosphere. Nevertheless, all compartments were dominated by Ensifer, the genus of rhizobia that forms root nodule symbiosis with M. truncatula, and additional shotgun metagenomic sequencing suggests that the nodulating Ensifer were not genetically distinguishable from those elsewhere in the plant. We also identify a handful of OTUs that are common in nodule tissues, which are likely colonized from the root endosphere.ConclusionsOur results demonstrate strong host filtering effects, with rhizospheres driven by soil origin and internal plant compartments driven by host genetics, and identify several key nodule-inhabiting taxa that coexist with rhizobia in the native range. Our results set the stage for future functional genetic experiments aimed at expanding our pairwise understanding of legume-rhizobium symbiosis toward a more mechanistic understanding of plant microbiomes.4yG9Be_5fuxNDH9Lv9Ven5Video

MiCROPe 2019 - emerging research priorities towards microbe-assisted crop production.

ABSTRACTThe miCROPe 2019 symposium, which took place from 2 to 5 December 2019 in Vienna, Austria, has unified researchers and industry from around the world to discuss opportunities, challenges and needs of microbe-assisted crop production. There is broad consensus that microorganisms—with their abilities to alleviate biotic and abiotic stresses and to improve plant nutrition—offer countless opportunities to enhance plant productivity and to ameliorate agricultural sustainability. However, microbe-assisted cultivation approaches face challenges that need to be addressed before a breakthrough of such technologies can be expected. Following up on the miCROPe symposium and a linked satellite workshop on breeding for beneficial plant–microbe interactions, we carved out research priorities towards successful implementation of microbiome knowledge for modern agriculture. These include (i) to solve context dependency for microbial inoculation approaches and (ii) to identify the genetic determinants to allow breeding for beneficial plant–microbiome interactions. With the combination of emerging third generation sequencing technologies and new causal research approaches, we now stand at the crossroad of utilising microbe-assisted crop production as a reliable and sustainable agronomic practice.

Bacterial communities in the plant phyllosphere harbour distinct responders to a broad-spectrum pesticide

Pesticide application can be accompanied by harmful non-target effects that affect humans, animals, as well as whole ecosystems. However, such effects remain mainly unaddressed in connection with microorganisms, and especially bacteria therein, which are essential for ecosystem functioning and host health. We analysed bacterial communities by sequencing 16S rRNA gene fragment amplicons following spray application of a broad-spectrum fungicide based on the active ingredient N-(3,5-dichlorophenyl) succinimide on Nicotiana tabacum L. leaves. The plant's phyllosphere was predominantly colonized by Proteobacteria, with Alphaproteobacteria accounting for up to 33.8% of the indigenous bacterial community. Bioinformatic analyses indicated that pesticide applications had an effect on the core microbiome as well as the rare microbiome. Moreover, the interference of the pesticide with phyllosphere bacteria was found to be selective. We have identified four positive responders including an ASV assigned to the genus Acinetobacter and 12 negative responders mainly assigned to bacterial genera known for beneficial plant-microbe interactions, including Stenotrophomonas, Sphingomonas, Flavobacterium and Serratia. Complementary inference of bacterial functioning on community level indicated that microbes with distinct stress response systems were likely enriched in the conducted treatments. The overall findings confirmed that pesticide treatments can induce measureable shifts in non-target bacterial communities colonizing the plant phyllosphere. They also indicate that potentially beneficial bacteria, which are known for their intrinsic association with plants, are among the most sensitive responders to the employed fungicide and thus highlight the importance of off-target studies in the context of the plant microbiome.

Reclamation with organic amendments and plants remodels the diversity and structure of bacterial community in ion-adsorption rare earth element mine tailings

The success of phytoremediation relies on beneficial plant-microbe interactions. However, the changes of soil microbiota, during the phytoremediation of ion-adsorption rare earth element (REE) mine tailings, are far from understood. The present study was conducted to reveal the co-occurrence patterns and key regulating factors of bacterial communities in ion-adsorption REE mine tailings with phytoremediation. A field experiment was conducted on an ion-adsorption REE mine tailing to test three phytoremediation strategies: (i) phytostabilization with grasses, (ii) phytostabilization with economic crops, and (iii) phytoextraction with REE hyperaccumulators. The bacterial community diversity, co-occurrence patterns, and the key regulating environmental factors in bulk and rhizospheric soils after 16 months of reclamation were studied by 16S Illumina high-throughput sequencing. The soils of plots with REEs-hyperaccumulators had significantly higher α-diversity than those planted with non-accumulators. After phytoremediation, the diversity and relative abundances of bacteria assisting in nutrient acquisition pronouncedly increased in the bulk soil of the reclaimed plots. Some plant growth–promoting bacteria (PGPB), e.g., Rhodanobacter, Streptomyces, and Bacillus, were enriched in the rhizospheric soil samples. Meanwhile, soil nutrient (total carbon and total nitrogen) concentration and REE availability were the most significant factors shaping the bacterial communities. Furthermore, special bacterial consortia of Bacilli took up the positions of the keystone species. The results revealed the co-occurrence patterns and key regulating factors of bacterial communities in ion-adsorption REE mine tailings, which will provide more crucial information for the optimization of the reclamation of REE tailings.

Comparative Analysis of Rhizosphere Microbiomes of Southern Highbush Blueberry (Vaccinium corymbosum L.), Darrow's Blueberry (V. darrowii Camp), and Rabbiteye Blueberry (V. virgatum Aiton).

Plants are inhabited by millions of parasitic, commensal, and mutualistic microorganisms that coexist in complex ecological communities, and profoundly affect the plant’s productivity, health, and capacity to cope with environmental stress. Therefore, a better understanding of the rhizosphere microbiome may open a yet untapped avenue for the rational exploitation of beneficial plant–microbe interactions in modern agriculture. Blueberries encompass several wild and cultivated species of shrubs of the genus Vaccinium that are native to North America. They are grown commercially for the production of fruits, which are considered a health food due to the rich content of minerals, trace elements, and phenolic compounds with antioxidant, antitumor, and anti-inflammatory properties. Despite a long history of breeding and extensive commercial use, remarkably little is known about the composition and function of the blueberry root microbiome. To address this gap, we employed molecular approaches to characterize and compare microbial communities inhabiting the roots of rabbiteye blueberry (Vaccinium virgatum), Darrow’s blueberry (Vaccinium darrowii), and southern highbush blueberry (SHB; an interspecific hybrid of Vaccinium corymbosum and V. darrowii). Our results revealed that these plant species share a common core rhizobiome, but at the same time differ significantly in the diversity, relative abundance, richness, and evenness of multiple groups of prokaryotic and eukaryotic microorganisms. Although the host signature effects were especially pronounced at the plant species level, we also observed genotype-level variations in the distribution of specific microbial taxa, which suggests that the assembly of the blueberry microbiome is shaped by the plant genotype and modifications associated with the domestication and breeding of members of the Vaccinium genus. We also demonstrated that the studied Vaccinium species differ in the abundance of beneficial rhizobacteria and ericoid mycorrhizal fungi, which play a vital role in their adaptation to soils with low pH and slow turnover of organic matter.

Arbuscular mycorrhizal fungi (AMF) influences growth and insect community dynamics in Sorghum-sudangrass (Sorghum x drummondii)

Beneficial plant–microbe interactions in the rhizosphere have been found to enhance plant growth and development. Arbuscular mycorrhizal fungi (AMF), a major group among these microbes, have been found to improve plant fitness through mycorrhizal symbiosis. Despite being well documented in various natural and domesticated study systems, few studies have examined whether AMF also has cascading effects on other traits, such as influencing insect community dynamics through attraction/repulsion of beneficial and harmful insects. To test this, we planted Sorghum-sudangrass (Sorghum x drummondii), a fast-growing annual grain/forage crop, either inoculated with commercial AMF mix or left as control in lab and field experiments. We hypothesized that AMF would enhance plant growth and influence the recruitment of insect herbivores and their natural enemies due to possible alterations in plant defense pathways. Our results suggest that while AMF-inoculated plants had significantly better germination, growth, and establishment; they also experienced a lower initial incidence of Spodoptera frugiperda, a major herbivore on Sorghum in the Lower Rio Grande Valley. In addition, our insect community trapping experiment revealed that AMF-inoculated plants attracted significantly more beneficial insects (predators and parasitoids) and a lower number of damaging herbivores. Taken together, our field and lab data show that AMF can not only positively influence plant growth traits but can also provide defenses against herbivores by selectively attracting beneficial insects and repelling herbivores, with implications for sustainable pest management strategies.

Effects of Soil Microbes on Functional Traits of Loblolly Pine (Pinus taeda) Seedling Families From Contrasting Climates.

Examining factors that influence seedling establishment is essential for predicting the impacts of climate change on tree species’ distributions. Seedlings originating from contrasting climates differentially express functional traits related to water and nutrient uptake and drought resistance that reflect their climate of origin and influence their responses to drought. Soil microbes may improve seedling establishment because they can enhance water and nutrient uptake and drought resistance. However, the relative influence of soil microbes on the expression of these functional traits between seedling families or populations from contrasting climates is unknown. To determine if soil microbes may differentially alter functional traits to enhance water and nutrient uptake and drought resistance between dry and wet families, seeds of loblolly pine families from the driest and wettest ends of its geographic range (dry, wet) were planted in sterilized sand (controls) or in sterilized sand inoculated with a soil microbial community (inoculated). Functional traits related to seedling establishment (germination), water and nutrient uptake and C allocation (root:shoot biomass ratio, root exudate concentration, leaf C:N, leaf N isotope composition (δ15N)), and drought resistance (turgor loss point, leaf carbon isotope composition (δ13C)) were measured. Then, plants were exposed to a drought treatment and possible shifts in photosynthetic performance were monitored using chlorophyll fluorescence. Inoculated plants exhibited significantly greater germination than controls regardless of family. The inoculation treatment significantly increased root:shoot biomass ratio in the wet family but not in the dry family, suggesting soil microbes alter functional traits that improve water and nutrient uptake more so in a family originating from a wetter climate than in a family originating from a drier climate. Microbial effects on photosynthetic performance during drought also differed between families, as photosynthetic performance of the dry inoculated group declined fastest. Regardless of treatment, the dry family exhibited a greater root:shoot biomass ratio, root exudate concentration, and leaf δ15N than the wet family. This indicates that the dry family allocated more resources belowground than the wet and the two family may have used different sources of plant available N, which may be related to their contrasting climates of origin and influence their drought resistance. Examination of variation in impacts of soil microbes on seedling physiology improves efforts to enhance seedling establishment and beneficial plant-microbe interactions under climate change.

Rapid Evolution of Plant-Bacterium Mutualism in the Rhizosphere

Plant roots are associated with a diverse microbial community that consists of antagonistic, commensal and mutualistic organisms. While beneficial plant-microbe interactions are important for plant performance, direct evidence for the evolution of rhizosphere mutualism remains elusive. Here we experimentally show that initially plant-antagonistic Pseudomonas protegens bacterium can rapidly evolve into a mutualist in the rhizosphere of Arabidopsis thaliana only within six plant growth cycles (6 months). Using an in vivo experimental evolution system, we observed that P. protegens rapidly diversified into distinct phenotypes, with some having clearly positive effects on the plant performance. Crucially, this evolution of plant facilitation was also accompanied with increased bacterial fitness indicative of a mutualistic interaction. At the genetic level, P. protegens mutualism was associated with mutations in the GacS/GacA two-component global regulator system, which is known to affect several aspects of bacterial physiology including carbon metabolism, host-microbe signalling, and downregulation of phytotoxic secondary metabolites. At the phenotypic level, the evolution of mutualism could be explained via two mechanisms: improved competition for root exudates and enhanced capacity for activating the root-specific transcription factor gene MYB72, which is essential for the production and excretion of the antimicrobial scopoletin for which the mutualists evolved enhanced tolerance. Together, these results show that plant-bacteria mutualism can rapidly evolve at an agriculturally relevant evolutionary timescale in the rhizosphere.

Genome-wide investigation of GRAM-domain containing genes in rice reveals their role in plant-rhizobacteria interactions and abiotic stress responses

A comprehensive genome-wide survey of GRAM-domain containing genes in rice identified total 64 genes which were grouped into six classes and were physically mapped onto different rice chromosomes. GRAM domain-containing genes showed total 8 segmental and 3 tandem duplications. Comparative physical mapping between rice OsGRAM and its orthologs in related C4-crops depicted evolutionary insights into this gene family. Expression analyses of OsGRAM genes in rice roots subjected to salt stress with or without Bacillus amyloliquefaciens (SN13) inoculation revealed significant differential expression patterns suggesting their crucial role in beneficial plant-rhizobacteria interactions under stress. Further, expression analyses of selected 15 candidate genes with ≥3.0-fold induction in salt + SN13 treated samples indicated their precise and overlapping expression patterns under various abiotic stresses and phytohormones at early (1 h) and late (24 h) durations which might be ultimately responsible for functional divergence and beneficial plant-microbe interactions. Furthermore, OsGRAM27 and OsGRAM47 could be considered as potential candidate genes for further functional characterization and application in crop improvement since these genes showed positive modulation in stress under the influence of SN13. This study provides new dimensions into the evolution and divergence of OsGRAM and their role in plant-rhizobacteria interactions that could be utilized for improving stress tolerance in crops.

Agricultural and Other Biotechnological Applications Resulting from Trophic Plant-Endophyte Interactions

Endophytic microbiota plays a role not only in supplying plants with the basic nutrients indispensable for their growth, but also helps them in the mechanisms of adaptation to various environmental stresses (i.e., salinity, drought), which is important in the aspect of crop yields. From the agricultural and biotechnological points of view, the knowledge of endophytes and their roles in increasing crop yields, plant resistance to diseases, and helping to survive environmental stress is extremely desirable. This paper reviews some of the beneficial plant–microbe interactions that might be potentially used in both agriculture (plant growth stimulation effect, adaptation of host organisms in salinity and drought conditions, and support of defense mechanisms in plants), and in biotechnology (bioactive metabolites, application of endophytes for bioremediation and biotransformation processes, and production of biofertilizers and biopreparations). Importantly, relatively recent reports on endophytes from the last 10 years are summarized in this paper.

Metabolites in the root exudates of groundnut change during interaction with plant growth promoting rhizobacteria in a strain-specific manner

Plant growth promoting rhizobacteria (PGPR) are extensively used as biofertilizers to improve the soil nutrition for a variety of crop plants. The plant-PGPR interaction, with special reference to chemical signalling molecules is not understood clearly, unlike other beneficial plant-microbe interactions. Chemo-attraction of a PGPR from soil microbial pool towards a plant could be dependent on some of the molecules in the plant root exudates (REs), similar to the beneficial association of legume-rhizobia. In this study, a few functional properties of PGPR like growth, chemotaxis, and biofilm formation by two PGPR strains viz., Bacillus sonorensis RS4 and Pseudomonas aeruginosa RP2 were assessed in the presence of groundnut REs. Functional properties of both the strains were significantly influenced by the REs in a strain-dependent manner. Metabolite profiling of the REs from PGPR-bacterized (RS4 or RP2) and non-bacterized seedlings was performed with GC–MS/MS after 12 and 24 days of growth. A total of 75 metabolites were detected in groundnut REs. Threonine and glyoxylic oxime acid were detected in RP2-bacterized REs, while serine, pentanoic acid, glucopyranoside, tartaric acid, and 2-pyrrolidinone were detected in REs of seedlings bacterized with RP2 and RS4. The results suggested that the PGPR induced distinct variations in the REs. Identification of the interaction-specific metabolites will be useful to develop effective PGPR based bio-formulations for better PGPR colonization and improving crop yields.

Transcriptional alterations reveal Bacillus amyloliquefaciens-rice cooperation under salt stress

The Bacillus amyloliquefaciens-SN13 and model crop rice (Oryza sativa) were chosen to understand the complex regulatory networks that govern plant-PGPR interaction under salt stress. During stress, inoculation with SN13 significantly increased biomass, relative water content, proline and total soluble sugar in rice while decreased lipid peroxidation and electrolyte leakage. Extensive alterations in gene expression were also observed in rice root transcriptome under stress in the presence of SN13. Rhizobacteria induced changes in expression of a considerable number of photosynthesis, hormone, and stress-responsive genes, in addition to cell-wall and lipid metabolism-related genes under salt stress as compared to salt stress or SN13 inoculation alone, indicating its potential role in reducing the harmful effects of salinity. To validate RNA-seq data, qRT-PCR was performed for selected differentially expressed genes representing various functional categories including metabolism, regulation, stress-response, and transporters. Results indicate qualitative and quantitative differences between roots responses to SN13 under stressed and unstressed conditions. Functional expressions of OsNAM and OsGRAM in yeast showed enhanced tolerance to various abiotic stresses, indicating crucial SN13-rice interaction in imparting beneficial effects under stress. This is first detailed report on understanding molecular mechanism underlying beneficial plant-microbe interaction in any economically important model crop plant under abiotic stress.

Discussion paper: Sustainable increase of crop production through improved technical strategies, breeding and adapted management – A European perspective

During the next decade it will be necessary to develop novel combinations of management strategies to sustainably increase crop production and soil resilience. Improving agricultural productivity, while conserving and enhancing biotic and abiotic resources, is an essential requirement to increase global food production on a sustainable basis. The role of farmers in increasing agricultural productivity growth sustainably will be crucial. Farmers are at the center of any process of change involving natural resources and for this reason they need to be encouraged and guided, through appropriate incentives and governance practices, to conserve natural ecosystems and their biodiversity, and minimize the negative impact agriculture can have on the environment. Farmers and stakeholders need to revise traditional approaches not as productive as the modern approaches but more friendly with natural and environmental ecosystems values as well as emerging novel tools and approaches addressing precise farming, organic amendments, lowered water consumption, integrated pest control and beneficial plant-microbe interactions. While practical solutions are developing, science based recommendations for crop rotations, breeding and harvest/postharvest strategies leading to environmentally sound and pollinator friendly production and better life in rural areas have to be provided.

Enrichment of Verrucomicrobia, Actinobacteria and Burkholderiales drives selection of bacterial community from soil by maize roots in a traditional milpa agroecosystem.

Milpas are rain-fed agroecosystems involving domesticated, semi-domesticated and tolerated plant species that combine maize with a large variety of other crop, tree or shrub species. Milpas are low input and low-tillage, yet highly productive agroecosystems, which have been maintained over millennia in indigenous communities in Mexico and other countries in Central America. Thus, milpas may retain ancient plant-microorganisms interactions, which could have been lost in modern high-tillage monocultures with large agrochemical input. In this work, we performed high-throughput 16S ribosomal DNA sequencing of soil adjacent to maize roots and bulk soil sampled at 30 cm from the base of the plants. We found that the bacterial communities of maize root soil had a lower alpha diversity, suggesting selection of microorganisms by maize-roots from the bulk-soil community. Beta diversity analysis confirmed that these environments harbor two distinct microbial communities; differences were driven by members of phyla Verrucomicrobia and Actinobacteria, as well as the order Burkholderiales (Betaproteobacteria), all of which had higher relative abundance in soil adjacent to the roots. Numerous studies have shown the influence of maize plants on bacterial communities found in soil attached tightly to the roots; here we further show that the influence of maize roots at milpas on bacterial communities is detectable even in plant-free soil collected nearby. We propose that members of Verrucomicrobia and other phyla found in the rhizosphere may establish beneficial plant-microbe interactions with maize roots in milpas, and propose to address their cultivation for future studies on ecology and potential use.

RiCRN1, a Crinkler Effector From the Arbuscular Mycorrhizal Fungus Rhizophagus irregularis, Functions in Arbuscule Development.

Arbuscular mycorrhizal (AM) symbiosis is one of the most prominent and beneficial plant–microbe interactions that facilitates mineral nutrition and confers tolerance to biotic and abiotic stresses. AM fungi colonize the root cortex and develop specialized structures called arbuscules where the nutrient exchange takes place. Arbuscule development is a highly controlled and coordinated process requiring the involvement of many plant proteins recruited at that interface. In contrast, much less is known about the fungal proteins involved in this process. Here, we have identified an AM fungal effector that participates in this developmental step of the symbiosis. RiCRN1 is a crinkler (CRN) effector that belongs to a subfamily of secreted CRN proteins from R. irregularis. CRNs have been so far only functionally characterized in pathogenic microbes and shown to participate in processes controlling plant cell death and immunity. RiCRN1 accumulates during symbiosis establishment parallel to MtPT4, the gene coding for an arbuscule-specific phosphate transporter. Expression in Nicotiana benthamiana leaves and in Medicago truncatula roots suggest that RiCRN1 is not involved in cell death processes. RiCRN1 dimerizes and localizes to nuclear bodies, suggesting that, similar to other CRNs, it functions in the plant nucleus. Downregulation of RiCRN1 using host-induced gene silencing led to an impairment of the symbiosis in M. truncatula and to a reduction of MtPT4, while ectopic expression of RiCRN1, surprisingly, led to a drastic reduction in arbuscule size that correlated with a decrease not only in MtPT4 but also in MtBCP1, a marker for initial stages of arbuscule development. Altogether, our results suggest that a tightly regulated expression in time and space of RiCRN1 is critical for symbiosis progression and for the proper initiation of arbuscule development.

Defining plant growth promoting rhizobacteria molecular and biochemical networks in beneficial plant-microbe interactions

Our knowledge of plant beneficial bacteria in the rhizosphere is rapidly expanding due to intense interest in utilizing these types of microbes in agriculture. Laboratory and field studies consistently document the growth, health and protective benefits conferred to plants by applying plant growth promoting rhizobacteria (PGPR). PGPR exert their influence on other species, including plants, in the rhizosphere by producing a wide array of extracellular molecules for communication and defense. The types of PGPR molecular products are characteristically diverse, and the mechanisms by which they are acting on the plant are only beginning to be understood. While plants may contribute to shape their microbiome, it is these bacterial products which induce beneficial responses in plants. PGPR extracellular products can directly stimulate plant genetic and molecular pathways, leading to increases in plant growth and induction of plant resistance and tolerance. This review will discuss known PGPR-derived molecules, and how these products are implicated in inducing plant beneficial outcomes through complex plant response mechanisms. In order to move PGPR research to the next level, it will be important to describe and document the genetic and molecular mechanisms employed in these interactions. In this way, we will be able to restructure and harness these mechanisms in a way that allows for broad-based applications in agriculture. A greater depth of understanding of how these PGPR molecules are acting on the plant will allow more effective development of rhizobacterial applications in the field.

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.

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.