Extracellular Vesicle-Driven Crosstalk between Legume Plants and Rhizobia: The Peribacteroid Space of Symbiosomes as a Protein Trafficking Interface.

Prokaryotes and eukaryotes secrete extracellular vesicles (EVs) into the surrounding milieu to preserve and transport elevated concentrations of biomolecules across long distances. EVs encapsulate metabolites, DNA, RNA, and proteins, whose abundance and composition fluctuate depending on environmental cues. EVs are involved in eukaryote-to-prokaryote communication owing to their ability to navigate different ecological niches and exchange molecular cargo between the two domains.

Isolation of Rhizobial Extracellular Membrane Vesicles from Bacteroids.

Extracellular-membrane vesicles (EMVs) are spherical buds of the extracellular membrane, commonly produced by Gram-negative bacteria, known to mediate intricate inter-kingdom communication. In this context, comprehensive research dissecting the role of EMVs in one of the most complex nature-occurring molecular dialogues, rhizobium-legume symbiosis, has been so far neglected. During the different stages of the symbiotic process, rhizobia and their host plants establish a very specific and controlled intercellular trafficking of signal molecules.

Isolation, Quantification, and Visualization of Extracellular Membrane Vesicles in Rhizobia Under Free-Living Conditions.

Rhizobia are a group of soil proteobacteria that are able to establish a symbiotic interaction with legumes. These bacteria are capable to fix atmospheric nitrogen into ammonia within specific plant root organs called nodules. The rhizobia-legume interaction is established by a complex molecular dialogue that starts with flavonoids exudated by the plant roots.

Virulence-Related Assays for Investigation of the Acidovorax citrulli-Cucurbitaceae Pathosystem.

Acidovorax citrulli is one of the most important pathogens of cucurbit crops, mainly melon and watermelon. Although A. citrulli is able to infect all aerial parts of the plant, fruits are highly sensitive to the bacterium.

A complex regulatory network governs the expression of symbiotic genes in HH103.

Introduction: The establishment of the rhizobium-legume nitrogen-fixing symbiosis relies on the interchange of molecular signals between the two symbionts. We have previously studied by RNA-seq the effect of the symbiotic regulators NodD1, SyrM, and TtsI on the expression of the symbiotic genes (the regulon) of HH103 upon treatment with the isoflavone genistein. In this work we have further investigated this regulatory network by incorporating new RNA-seq data of HH103 mutants in two other regulatory genes, and .

Natural variation in a short region of the Acidovorax citrulli type III-secreted effector AopW1 is associated with differences in cytotoxicity and host adaptation.

Bacterial fruit blotch, caused by Acidovorax citrulli, is a serious disease of melon and watermelon. The strains of the pathogen belong to two major genetic groups: group I strains are strongly associated with melon, while group II strains are more aggressive on watermelon. A.

Membrane vesicle engineering with "à la carte" bacterial-immunogenic molecules for organism-free plant vaccination.

The United Nations heralds a world population exponential increase exceeding 9.7 billion by 2050. This poses the challenge of covering the nutritional needs of an overpopulated world by the hand of preserving the environment.

Non-Ionic Osmotic Stress Induces the Biosynthesis of Nodulation Factors and Affects Other Symbiotic Traits in HH103.

(1) Background: Some rhizobia, such as CIAT 899, activate nodulation genes when grown under osmotic stress. This work aims to determine whether this phenomenon also takes place in HH103. (2) Methods: HH103 was grown with and without 400 mM mannitol.

The Rhizobium tropici CIAT 899 NodD2 protein promotes symbiosis and extends rhizobial nodulation range by constitutive nodulation factor synthesis.

In the symbiotic associations between rhizobia and legumes, the NodD regulators orchestrate the transcription of the specific nodulation genes. This set of genes is involved in the synthesis of nodulation factors, which are responsible for initiating the nodulation process. Rhizobium tropici CIAT 899 is the most successful symbiont of Phaseolus vulgaris and can nodulate a variety of legumes.

Knock-down of phosphoenolpyruvate carboxylase 3 negatively impacts growth, productivity, and responses to salt stress in sorghum (Sorghum bicolor L.).

Phosphoenolpyruvate carboxylase (PEPC) is a carboxylating enzyme with important roles in plant metabolism. Most studies in C plants have focused on photosynthetic PEPC, but less is known about non-photosynthetic PEPC isozymes, especially with respect to their physiological functions. In this work, we analyzed the precise roles of the sorghum (Sorghum bicolor) PPC3 isozyme by the use of knock-down lines with the SbPPC3 gene silenced (Ppc3 lines).

The Gene of HH103 Restores Nodulation Capacity on Bean in a CIAT 899 / Mutant, but the Secondary Symbiotic Regulators , or Prevent HH103 to Nodulate with This Legume.

Rhizobial NodD proteins and appropriate flavonoids induce rhizobial nodulation gene expression. In this study, we show that the gene of HH103, but not the gene, can restore the nodulation capacity of a double / mutant of CIAT 899 in bean plants (). HH103 only induces pseudonodules in beans.

Secretion systems play a critical role in resistance to predation by Bdellovibrio bacteriovorus.

Bdellovibrio bacteriovorus, a Gram-negative predatory bacterium belonging to the Bdellovibrio and like organisms (BALOs), predates on Gram-negative bacteria. BALO strains differ in prey range but so far, the genetic basis of resistance against BALO predation is hardly understood. We developed a loss-of-function approach to screen for sensitive mutants in a library of strain M6, a predation-resistant strain of the plant pathogen Acidovorax citrulli.

One door closes, another opens: when nodulation impairment with natural hosts extends rhizobial host-range.

The rhizobium-legume symbiosis is the best-understood plant-microbe association. The high degree of specificity observed in this relationship is supported by a complex exchange of signals between the two components of the symbiosis. Findings reported in last years indicate that multiple molecular mechanisms, such as the production of a particular set of nodulation factors at a very specific concentration or a suitable arsenal of effectors secreted through the type III secretion system, have been adjusted during evolution to ensure and optimize the recognition of specific rhizobial strains by its legume host.

OnfD, an AraC-Type Transcriptional Regulator Encoded by CIAT 899 and Involved in Nod Factor Synthesis and Symbiosis.

CIAT 899 is a broad-host-range rhizobial strain that establishes symbiotic interactions with legumes and tolerates different environmental stresses such as heat, acidity, or salinity. This rhizobial strain produces a wide variety of symbiotically active nodulation factors (NF) induced not only by the presence of plant-released flavonoids but also under osmotic stress conditions through the LysR-type transcriptional regulators NodD1 (flavonoids) and NodD2 (osmotic stress). However, the activation of NodD2 under high-osmotic-stress conditions remains elusive.

The Sinorhizobium fredii HH103 type III secretion system effector NopC blocks nodulation with Lotus japonicus Gifu.

The broad-host-range bacterium Sinorhizobium fredii HH103 cannot nodulate the model legume Lotus japonicus Gifu. This bacterium possesses a type III secretion system (T3SS), a specialized secretion apparatus used to deliver effector proteins (T3Es) into the host cell cytosol to alter host signaling and/or suppress host defence responses to promote infection. However, some of these T3Es are recognized by specific plant receptors and hence trigger a strong defence response to block infection.

Deciphering the Symbiotic Significance of Quorum Sensing Systems of HH103.

Quorum sensing (QS) is a bacterial cell-to-cell signaling mechanism that collectively regulates and synchronizes behaviors by means of small diffusible chemical molecules. In rhizobia, QS systems usually relies on the synthesis and detection of -acyl-homoserine lactones (AHLs). In the model bacterium functions regulated by the QS systems TraI-TraR and SinI-SinR(-ExpR) include plasmid transfer, production of surface polysaccharides, motility, growth rate and nodulation.

Show me your secret(ed) weapons: a multifaceted approach reveals a wide arsenal of type III-secreted effectors in the cucurbit pathogenic bacterium Acidovorax citrulli and novel effectors in the Acidovorax genus.

The cucurbit pathogenic bacterium Acidovorax citrulli requires a functional type III secretion system (T3SS) for pathogenicity. In this bacterium, as with Xanthomonas and Ralstonia spp., an AraC-type transcriptional regulator, HrpX, regulates expression of genes encoding T3SS components and type III-secreted effectors (T3Es).

Complete Assembly of the Genome of an Strain Reveals a Naturally Occurring Plasmid in This Species.

is the causal agent of bacterial fruit blotch (BFB), a serious threat to cucurbit crop production worldwide. Based on genetic and phenotypic properties, strains are divided into two major groups: group I strains have been generally isolated from melon and other non-watermelon cucurbits, while group II strains are closely associated with watermelon. In a previous study, we reported the genome of the group I model strain, M6.

Osmotic stress activates nif and fix genes and induces the Rhizobium tropici CIAT 899 Nod factor production via NodD2 by up-regulation of the nodA2 operon and the nodA3 gene.

The symbiosis between rhizobia and legumes is characterized by a complex molecular dialogue in which the bacterial NodD protein plays a major role due to its capacity to activate the expression of the nodulation genes in the presence of appropiate flavonoids. These genes are involved in the synthesis of molecules, the nodulation factors (NF), responsible for launching the nodulation process. Rhizobium tropici CIAT 899, a rhizobial strain that nodulates Phaseolus vulgaris, is characterized by its tolerance to multiple environmental stresses such as high temperatures, acidity or elevated osmolarity.

Sinorhizobium fredii HH103 nolR and nodD2 mutants gain capacity for infection thread invasion of Lotus japonicus Gifu and Lotus burttii.

Sinorhizobium fredii HH103 Rif , a broad-host-range rhizobial strain, forms ineffective nodules with Lotus japonicus but induces nitrogen-fixing nodules in Lotus burttii roots that are infected by intercellular entry. Here we show that HH103 Rif nolR or nodD2 mutants gain the ability to induce infection thread formation and to form nitrogen-fixing nodules in L. japonicus Gifu.

Transcriptomic Studies of the Effect of nod Gene-Inducing Molecules in Rhizobia: Different Weapons, One Purpose.

Simultaneous quantification of transcripts of the whole bacterial genome allows the analysis of the global transcriptional response under changing conditions. RNA-seq and microarrays are the most used techniques to measure these transcriptomic changes, and both complement each other in transcriptome profiling. In this review, we exhaustively compiled the symbiosis-related transcriptomic reports (microarrays and RNA sequencing) carried out hitherto in rhizobia.

The Rhizobium tropici CIAT 899 NodD2 protein regulates the production of Nod factors under salt stress in a flavonoid-independent manner.

In the symbiotic associations between rhizobia and legumes, NodD promotes the expression of the nodulation genes in the presence of appropriate flavonoids. This set of genes is implied in the synthesis of Nodulation factors, which are responsible for launching the nodulation process. Rhizobium tropici CIAT 899 is the most successful symbiont of Phaseolus vulgaris and can nodulate a variety of legumes.

The Sinorhizobium (Ensifer) fredii HH103 Nodulation Outer Protein NopI Is a Determinant for Efficient Nodulation of Soybean and Cowpea Plants.

The type III secretion system (T3SS) is a specialized secretion apparatus that is commonly used by many plant and animal pathogenic bacteria to deliver proteins, termed effectors, to the interior of the host cells. These effectors suppress host defenses and interfere with signal transduction pathways to promote infection. Some rhizobial strains possess a functional T3SS, which is involved in the suppression of host defense responses, host range determination, and symbiotic efficiency.

NrcR, a New Transcriptional Regulator of Rhizobium tropici CIAT 899 Involved in the Legume Root-Nodule Symbiosis.

The establishment of nitrogen-fixing rhizobium-legume symbioses requires a highly complex cascade of events. In this molecular dialogue the bacterial NodD transcriptional regulators in conjunction with plant inducers, mostly flavonoids, are responsible for the biosynthesis and secretion of Nod factors which are key molecules for successful nodulation. Other transcriptional regulators related to the symbiotic process have been identified in rhizobial genomes, including negative regulators such as NolR.

RNA-seq analysis of the Rhizobium tropici CIAT 899 transcriptome shows similarities in the activation patterns of symbiotic genes in the presence of apigenin and salt.

Background: Rhizobium tropici strain CIAT 899 establishes effective symbioses with several legume species, including Phaseolus vulgaris and Leucaena leucocephala. This bacterium synthesizes a large variety of nodulation factors in response to nod-gene inducing flavonoids and, surprisingly, also under salt stress conditions. The aim of this study was to identify differentially expressed genes in the presence of both inducer molecules, and analyze the promoter regions located upstream of these genes.