Simultaneous Impact of Rhizobacteria Inoculation and Leaf-Chewing Insect Herbivory on Essential Oil Production and VOC Emissions in Ocimum basilicum.

Inoculation with rhizobacteria and feeding by herbivores, two types of abiotic stress, have been shown to increase the production of secondary metabolites in plants as part of the defense response. This study explored the simultaneous effects of inoculation with Bacillus amyloliquefaciens GB03 (a PGPR species) and herbivory by third-instar Spodoptera frugiperda larvae on essential oil (EO) yield and volatile organic compound (VOC) emissions in Ocimum basilicum plants. The density of glandular trichomes was also examined, given that they are linked to EO production and VOC emission. Herbivory increased EO content, but inoculation on its own did not. When combined, however, the two treatments led to a 10-fold rise in EO content with respect to non-inoculated plants. VOC emissions did not significantly differ between inoculated and non-inoculated plants, but they doubled in plants chewed by the larvae with respect to their undamaged counterparts. Interestingly, no changes were observed in VOC emissions when the treatments were tested together. In short, the two biotic stressors elicited differing plant defense responses, mainly when EO was concerned. PGPR did not stimulate EO production, while herbivory significantly enhanced it and increased VOC emissions. The combined treatment acted synergistically, and in this case, PGPR inoculation may have had a priming effect that amplified plant response to herbivory. Peltate trichome density was higher in inoculated plants, those damaged by larvae, and those subjected to the combination of both treatments. The findings highlight the intricate nature of plant defense mechanisms against various stressors and hint at a potential strategy to produce essential oil through the combined application of the two stressors tested here.

Complete genome sequence of Bradyrhizobium sp. 62B, a native nitrogen-fixing rhizobium isolated from peanut nodules.

We present the complete genome sequence of Bradyrhizobium sp. 62B, a strain isolated from the root nodules of peanut plants that grow in central Argentina. The genome consists of 8.15 Mbp, distributed into a chromosome of 7.29 Mbp and a plasmid of 0.86 Mbp.

Exploring the Differential Impact of Salt Stress on Root Colonization Adaptation Mechanisms in Plant Growth-Promoting Rhizobacteria.

Salinity inhibits plant growth by affecting physiological processes, but soil microorganisms like plant growth-promoting rhizobacteria (PGPR) can alleviate abiotic stress and enhance crop productivity. However, it should be noted that rhizobacteria employ different approaches to deal with salt stress conditions and successfully colonize roots. The objective of this study was to investigate the effect of salt stress on bacterial survival mechanisms such as mobility, biofilm formation, and the autoaggregation capacity of three plant growth-promoting strains: Pseudomonas putida SJ04, Pseudomonas simiae WCS417r, and Bacillus amyloliquefaciens GB03. These strains were grown in diluted LB medium supplemented with 0, 100, 200, or 300 mM NaCl. Swimming and swarming mobility were evaluated in media supplemented with 0.3 and 0.5% agar, respectively. Biofilm formation capacity was quantified using the crystal violet method, and the autoaggregation capacity was measured spectrophotometrically. In addition, we evaluated in vitro the capacity of the strains to ameliorate the effects of saline stress in Mentha piperita. The study found that the GB03 strain exhibited enhanced swarming mobility when the salt concentration in the medium increased, resulting in a two-fold increase in the halo diameter at 300 mM. However, high concentrations of NaCl did not affect the swimming mobility. In contrast, swimming motility was reduced in WCS417r and SJ04 under salt stress. On the other hand, exposure to 300 mM NaCl resulted in a 180% increase in biofilm formation and a 30% rise in the percentage of autoaggregation in WCS417r. Conversely, the autoaggregation percentage of the strains SJ04 and GB03 remained unaffected by saline stress. However, for GB03, biofilm formation decreased by 80% at 300 mM. Simultaneously, inoculation with the three evaluated strains alleviated the detrimental effects of salinity on plant growth. Under 150 mM salt stress, all strains showed increased fresh weight, with GB03 and WCS417r improving by 40% and SJ04 exhibiting the most remarkable effect with a 70% rise compared to non-inoculated plants. Despite their different strategies for mitigating salt stress, the application of these strains presents a promising strategy for effectively mitigating the negative consequences of salt stress on plant cultivation.

Genome sequence of Mesorhizobium mediterraneum strain R31, a nitrogen-fixing rhizobium used as an inoculant for chickpea in Argentina.

Here, we report the complete genome sequence of Mesorhizobium mediterraneum R31, a rhizobial strain recommended and used as a commercial inoculant for chickpea in Argentina. The genome consists of 7.25 Mb, distributed into four circular replicons: a chromosome of 6.72 Mbp and three plasmids of 0.29, 0.17, and 0.07 Mbp.

Biofilm-Forming Ability of Phytopathogenic Bacteria: A Review of its Involvement in Plant Stress.

Phytopathogenic bacteria not only affect crop yield and quality but also the environment. Understanding the mechanisms involved in their survival is essential to develop new strategies to control plant disease. One such mechanism is the formation of biofilms; i.e., microbial communities within a three-dimensional structure that offers adaptive advantages, such as protection against unfavorable environmental conditions. Biofilm-producing phytopathogenic bacteria are difficult to manage. They colonize the intercellular spaces and the vascular system of the host plants and cause a wide range of symptoms such as necrosis, wilting, leaf spots, blight, soft rot, and hyperplasia. This review summarizes up-to-date information about saline and drought stress in plants (abiotic stress) and then goes on to focus on the biotic stress produced by biofilm-forming phytopathogenic bacteria, which are responsible for serious disease in many crops. Their characteristics, pathogenesis, virulence factors, systems of cellular communication, and the molecules implicated in the regulation of these processes are all covered.

Antifungal Activity and Alleviation of Salt Stress by Volatile Organic Compounds of Native Pseudomonas Obtained from Mentha piperita.

As salt stress has a negative impact on plant growth and crop yield, it is very important to identify and develop any available biotechnology which can improve the salt tolerance of plants. Inoculation with plant-growth-promoting rhizobacteria (PGPR) is a proven environmentally friendly biotechnological resource for increasing the salt stress tolerance of plants and has a potential in-field application. In addition, bacterial volatile organic compounds (mVOCs) are signal molecules that may have beneficial roles in the soil-plant-microbiome ecosystem. We investigated the effects of mVOCs emitted by Pseudomona putida SJ46 and SJ04 on Mentha piperita grown under different levels of NaCl stress by evaluating their growth-promoting potential and capacity to increase salt tolerance effects. Furthermore, we evaluated under control and salt stress conditions the biocontrol ability of VOCs emitted by both these strains to inhibit the growth of Alternaria alternata and Sclerotium rolfsii. The VOCs emitted by both strains under control conditions did not lead to an significant improvement in peppermint growth. However, under salt stress conditions (75 or 100 mM NaCl), an amelioration of its physiological status was observed, with this effect being greater at 100 mM NaCl. This led to an enhancement of the number of leaves and nodes and, increased the shoot fresh and root dry weight by approximately twice in relation to control stressed plants. Moreover, the VOCs released by the two bacteria grown in control or saline media showed a significant reduction in the mycelial growth of A. alternata. In contrast, S. rolfsii growth was reduced 40% by the mVOCs released only under control conditions, with no effects being observed under salt stress. We also explored the composition of the bacterial volatile profiles by means of a solid-phase microextraction/gas chromatography-mass spectrometry (SPME/GC-MS) analysis. From the headspace of SJ46, three VOCs were identified: n-octanol, decane and tetradecane. The emission of SJ04 had the same chromatographic profile, with the addition of two more compounds: 1-(N-phenyl carbamyl)-2-morpholino cyclohexene and tridecane. Only compounds that were not present in the headspace of the control groups were recorded. The salt stress conditions where the bacteria were grown did not qualitatively modify the mVOC emissions. Taken together, our results suggest that plant-associated rhizobacterial VOCs play a potentially important role in modulating plant salt tolerance and reducing fungal growth. Thus, biological resources represent novel tools for counteracting the deleterious effects of salt stress and have the potential to be exploited in sustainable agriculture. Nevertheless, future studies are necessary to investigate technological improvements for bacterial VOC application under greenhouse and open field conditions.

Whole-Genome Sequence of Burkholderia ambifaria Strain Q53, a Potential Plant Growth Promoter Isolated from the Rhizosphere of Peanut.

We report the complete genome sequence of Burkholderia ambifaria strain Q53, an environmental rhizobacterium isolated from the rhizosphere of peanut plants. The genome consists of 7.4 Mbp distributed into three circular chromosomes and was determined using a hybrid long- and short-read assembly approach.

A practical method to investigate the effect of volatile organic compounds emitted by rhizobacteria on plant growth under conditions of salt stress.

Salinity is one of the causes that limit crop production. Plant Growth Promoting Rhizobacteria (PGPR) are beneficial soil bacteria that play a significant role in promoting plant growth. These microorganisms can produce their effect through the emission of Volatile Organic Compounds (VOCs). Most of the research to study the effects of microbial VOCs on plant growth has been carried out under controlled conditions using partitioned Petri dishes. In this article, we describe an alternative method that has the advantage of allowing long-term trials, being able to let the plant have a greater development in growth and height, without space limitation. In the proposed method, M. piperita were planted in glass jars containing Murashige and Skoog solid media, with a small glass vial containing Hoagland media inserted into the jar. This small vial was inoculated with the specified bacterium and served as the source of bacterial volatiles. This way plants were exposed to mVOCs without having any physical contact with the rhizobacteria.•The procedure allows studying the effect of microbial VOCs on plant growth.•It also allows longer trials, being able to let the plant develop more without space limitation.

Coaggregative interactions between rhizobacteria are promoted by exopolysaccharides from Sinorhizobium meliloti.

Bacterial surface components and extracellular compounds such as exopolysaccharides (EPSs) are crucial for interactions between cells, tolerance to different types of stress, and host colonization. Sinorhizobium meliloti produces two EPSs: Succinoglycan (EPS I), which is involved in the establishment of symbiosis with Medicago sativa, and galactoglucan (EPS II), associated with biofilm formation and the promotion of aggregation. Here, we aimed to assess their role in aggregative interactions between cells of the same strain of a given species (auto-aggregation), and between genetically different strains of the same or different species (intra- or intergeneric coaggregation). To do this, we used S. meliloti mutants which are defective in the production of EPS I, EPS II, or both. Macroscopic and microscopic coaggregation tests were performed with combinations or pairs of different bacterial strains. The EPS II-producing strains were more capable of coaggregation than those that cannot produce EPS II. This was true both for coaggregations between different S. meliloti strains, and between S. meliloti and other common rhizobacteria of agricultural relevance, such as Pseudomonas fluorescens and Azospirillum brasilense. The exogenous addition of EPS II strongly promoted coaggregation, thus confirming the polymer's importance for this phenotype. EPS II may therefore be a key factor in events of physiological significance for environmental survival, such as aggregative interactions and biofilm development. Furthermore, it might be a connecting molecule with relevant properties at an ecological, biotechnological, and agricultural level.

Ascochyta Blight in Chickpea: An Update.

Chickpea (Cicer arietinum L.), one of the most cultivated legumes worldwide, is crucial for the economy of several countries and a valuable source of nutrients. Yields may be severely affected by Ascochyta blight, a disease caused by the fungus Ascochyta rabiei. Molecular and pathological studies have not yet managed to establish its pathogenesis, since it is highly variable. Similarly, much remains to be elucidated about plant defense mechanisms against the pathogen. Further knowledge of these two aspects is fundamental for the development of tools and strategies to protect the crop. This review summarizes up-to-date information on the disease's pathogenesis, symptomatology, and geographical distribution, as well as on the environmental factors that favor infection, host defense mechanisms, and resistant chickpea genotypes. It also outlines existing practices for integrated blight management.

Complete Genome Sequence of Mesorhizobium ciceri Strain R30, a Rhizobium Used as a Commercial Inoculant for Chickpea in Argentina.

We report the complete genome sequence of Mesorhizobium ciceri strain R30, a rhizobium strain recommended and used as a commercial inoculant for chickpea in Argentina. The genome consists of almost 7 Mb, distributed into two circular replicons: a chromosome of 6.49 Mb and a plasmid of 0.46 Mb.

Complete Genome Sequence of Bradyrhizobium sp. Strain C-145, a Nitrogen-Fixing Rhizobacterium Used as a Peanut Inoculant in Argentina.

We present the complete genome sequence of Bradyrhizobium sp. strain C-145, one of the most widely used nitrogen-fixing rhizobacteria for inoculating peanut crops in Argentina. The genome consists of 9.53 Mbp in a single circular chromosome and was determined using a hybrid long- and short-read assembly approach.

Beneficial rhizobacteria inoculation on Ocimum basilicum reduces the growth performance and nutritional value of Spodoptera frugiperda.

BACKGROUND: Plant growth-promoting rhizobacteria (PGPR) has a significant role in plant-insect interaction. However, the extent of their impact on insects is still not well understood. This investigation was designed to evaluate the role of inoculation with Bacillus amyloliquefaciens GB03 on sweet basil (Ocimum basilucum L.) in the development and nutritional parameters of Spodoptera frugiperda. In addition, the feeding preferences on inoculated and non-inoculated plants were assessed. RESULTS: Spodoptera frugiperda larvae reared with inoculated sweet basil leaves had a strong negative effect on the development of the insect, resulting in lower larval and pupal weights, and a longer period for larval-adult development. Moreover, adult emergence was reduced, but the relative consumption rate (RCR) value was unaffected, thereby revealing no alteration of the palatability. Growth rate and nutritional indicators, such as the efficiency of conversion of ingested food (ECI) and the efficiency of conversion of digested food (ECD), were reduced in larvae reared from treated plants. In the choice test, larvae avoided feeding on inoculated leaves. CONCLUSION: The higher occurrence of secondary metabolites in inoculated plants could have been the reason for the reduction of the plant nutritional rate and also for the food selection, since it has been previously reported that GB03 inoculated sweet basil increased the essential oil yield. Therefore, PGPR inoculation could be used as a growth promoter, making it a promising candidate for plant protection programs against insects in aromatic plant production. © 2021 Society of Chemical Industry.

A simple method to determine antioxidant status in aromatic plants subjected to drought stress.

Drought is a major environmental stress factor that affects the growth and development of plants. All plants have to maintain the reactive oxygen species within certain levels for normal cellular homeostasis by means of their antioxidant systems, which can be classified as enzymatic and non-enzymatic. Plants under drought stress generate an excess production of reactive oxygen species. At high concentrations, this can be detrimental by producing damage to the protein structures and inhibiting enzymes, as well as oxidizing macromolecules, which may eventually lead to cell death. There has been increasing attention paid to the antioxidant capacity of aromatic/medicinal plants, with a high antioxidant content having been reported in some plant extracts, such as in Mentha piperita (peppermint). Peppermint plants cultivated under drought stress also present high levels of phenolic compounds, peroxidase enzyme activity and lipid peroxidation of membranes. A simple and inexpensive laboratory class is proposed for teaching some mechanisms that plants have evolved to avoid reactive oxygen species damage. The series of lab experiments described is aimed at demonstrating the antioxidant status in aromatic plants subjected to drought stress, by measuring total phenolic compound content (non-enzymatic antioxidant compound), peroxidase activity (enzymatic antioxidant) and malondialdehyde, as convenient biomarkers for lipid peroxidation. The proposed class will be carried out by undergraduate students of the advanced biochemistry course, as part of our biology and agronomy studies. The experiment presented is intended to be used as a vehicle to emphasize the concepts that students have learned in their lectures. This lab exercise to be carried out by the students has dual goals: to apply a methodology only learned superficially on previous courses, and also to increase their understanding of how plants developed resistance mechanisms in order to tolerate drought stress.

Crystal structures of peanut lectin in the presence of synthetic ß-N- and ß-S-galactosides disclose evidence for the recognition of different glycomimetic ligands.

Carbohydrate-lectin interactions are involved in important cellular recognition processes, including viral and bacterial infections, inflammation and tumor metastasis. Hence, structural studies of lectin-synthetic glycan complexes are essential for understanding lectin-recognition processes and for the further design of promising chemotherapeutics that interfere with sugar-lectin interactions. Plant lectins are excellent models for the study of the molecular-recognition process. Among them, peanut lectin (PNA) is highly relevant in the field of glycobiology because of its specificity for ß-galactosides, showing high affinity towards the Thomsen-Friedenreich antigen, a well known tumor-associated carbohydrate antigen. Given this specificity, PNA is one of the most frequently used molecular probes for the recognition of tumor cell-surface O-glycans. Thus, it has been extensively used in glycobiology for inhibition studies with a variety of ß-galactoside and ß-lactoside ligands. Here, crystal structures of PNA are reported in complex with six novel synthetic hydrolytically stable ß-N- and ß-S-galactosides. These complexes disclosed key molecular-binding interactions of the different sugars with PNA at the atomic level, revealing the roles of specific water molecules in protein-ligand recognition. Furthermore, binding-affinity studies by isothermal titration calorimetry showed dissociation-constant values in the micromolar range, as well as a positive multivalency effect in terms of affinity in the case of the divalent compounds. Taken together, this work provides a qualitative structural rationale for the upcoming synthesis of optimized glycoclusters designed for the study of lectin-mediated biological processes. The understanding of the recognition of ß-N- and ß-S-galactosides by PNA represents a benchmark in protein-carbohydrate interactions since they are novel synthetic ligands that do not belong to the family of O-linked glycosides.

Evaluation of cellular safety and the chemical composition of the peanut (Arachis hypogaea L.) ethanolic extracts.

Arachis hypogaea L. (Leguminosae) is distributed in tropical and subtropical areas. Peanut has high nutritional and commercial value. Scientific research showed that peanut has biological properties such as anticancer, antioxidant, antiinflammatory. However, it is necessary to know if consumption of peanut, either as food or as a phytopharmaceutical implies a health risk. The aim was to evaluate cytotoxicity and genotoxicity of ethanolic extracts from A. hypogaea. Also, chemical characterization of these extracts was performed. Cytotoxicity was evaluated by MTT and Neutral Red Uptake (NRU) assays on Vero cells. Genotoxicity was studied by Micronuclei and comet assays on Balb/C mice. Qualitative and quantitative chemical analysis of extracts were performed. Results showed that extracts have low cytotoxicity. Tegument ethanolic extract (TEE) and Seed ethanolic extract (SEE) were not genotoxic. The treatments with TEE at 250 mg/kg and SEE at 2000 mg/kg revealed (highest concentrations evaluated) some toxicity on blood marrow cells of mice. Chemical characterization indicated that TEE had 74.33 ± 1.10 mg GAE/g of dried extract and SEE had 15.05 ± 0.06 mg GAE/g of dried extract of total phenolic content. Also, proanthocyanidins (O.D. at 550 nm 1.39 ± 0.15) and caffeic acid (2.46%) were identified in TEE. While, linoleic acid (58.84%) oleic acid (11.31%) and palmitic acid (8.37%) were major compounds of SEE. In conclusion, peanut consumption is safe at concentrations recommended for healthy uses, such as nutrition, and phytomedicine.

Exopolysaccharide II Is Relevant for the Survival of Sinorhizobium meliloti under Water Deficiency and Salinity Stress.

Sinorhizobium meliloti is a soil bacterium of great agricultural importance because of its ability to fix atmospheric nitrogen in symbiotic association with alfalfa (Medicago sativa) roots. We looked into the involvement of exopolysaccharides (EPS) in its survival when exposed to different environmental stressors, as well as in bacteria-bacteria and bacteria-substrate interactions. The strains used were wild-type Rm8530 and two strains that are defective in the biosynthesis of EPS II: wild-type Rm1021, which has a non-functional expR locus, and mutant Rm8530 expA. Under stress by water deficiency, Rm8530 remained viable and increased in number, whereas Rm1021 and Rm8530 expA did not. These differences could be due to Rm8530's ability to produce EPS II. Survival experiments under saline stress showed that viability was reduced for Rm1021 but not for Rm8530 or Rm8530 expA, which suggests the existence of some regulating mechanism dependent on a functional expR that is absent in Rm1021. The results of salinity-induced stress assays regarding biofilm-forming capacity (BFC) and autoaggregation indicated the protective role of EPS II. As a whole, our observations demonstrate that EPS play major roles in rhizobacterial survival.

Impact of Soil Rhizobacteria Inoculation and Leaf-Chewing Insect Herbivory on Mentha piperita Leaf Secondary Metabolites.

Secondary metabolites commonly play important physiological roles in plants and can be modified quantitatively and qualitatively by exposure to biotic and abiotic interactions. Plant growth promoting rhizobacteria (PGPR) and herbivory induce systemic resistance. In the present study, we analyzed the induction of secondary metabolites in peppermint plants in response to chewing insect herbivory on PGPR-inoculated Mentha piperita plants. The secondary metabolites of M. piperita plants were increased when plants were inoculated with PGPR and also exposed to caterpillar herbivory. It was found that the total essential oil yield in inoculated plants with insect damage was ~2.6-fold higher than in controls. The yield was similar to that of plants either damaged by insects or inoculated, indicating that there was no synergism. The same trend was observed for phenolic compounds. In contrast, VOC emissions were significantly higher in plants infested by insects, independent of whether they were inoculated. Insect damaged plants had 5.5 times higher monoterpene emissions than control plants, and ~ 2-fold higher emissions than on PGPR-inoculated plants without insects. To gain a better understanding of how herbivory on PGPR-inoculated plants can cause an increase in secondary metabolites of peppermint, we examined changes in plant defense hormones in inoculated plants after herbivory. We found that the combination of both treatments increased the endogenous jasmonic and salicylic acid levels to the same extent as in plants only inoculated or only insect-damaged. Because different interactions can alter the phytochemistry of plants such as M. piperita, this topic is both ecologically and economically relevant.

Proline metabolic dynamics and implications in drought tolerance of peanut plants.

Proline accumulation and metabolism are associated with mechanisms of abiotic stress avoidance in plants. Proline accumulation generally improves osmotic stress tolerance whereas proline metabolism can have varying effects from ATP generation to the formation of reactive oxygen species. To further understand the roles of proline in stress protection, two peanut cultivars with contrasting tolerance to drought were examined by transcriptional and biochemical analyses during water stress. Plants exposed to polyethylene glycol had diminished relative water content and increased proline content; while, only the drought sensitive plants, cultivar Granoleico, showed lipid oxidative damage (measured as thiobarbituric acid reactive substances). The expression of proline biosynthesis genes (P5CS1, P5CS2a, P5CS2b, P5CR) was increased in both cultivars upon exposure to water stress. However, the relative expression of proline catabolism genes (ProDH1, ProDH2) was increased only in the sensitive cultivar during stress. Exogenous addition of proline and the proline analogue thiazolidine-4-carboxylic acid (T4C), both substrates of proline dehydrogenase, was also used to exacerbate and identify plant responses. Pretreatment of plants with T4C induced unique changes in the drought tolerant EC-98 cultivar such as higher mRNA levels of proline biosynthetic and catabolic ProDH genes, even in the absence of water stress. The increased levels of ProDH gene expression, potentially associated with higher T4C conversion to cysteine, may contribute to the tolerant phenotype.

Exopolysaccharide production in Ensifer meliloti laboratory and native strains and their effects on alfalfa inoculation.

Bacterial surface molecules have an important role in the rhizobia-legume symbiosis. Ensifer meliloti (previously, Sinorhizobium meliloti), a symbiotic Gram-negative rhizobacterium, produces two different exopolysaccharides (EPSs), termed EPS I (succinoglycan) and EPS II (galactoglucan), with different functions in the symbiotic process. Accordingly, we undertook a study comparing the potential differences in alfalfa nodulation by E. meliloti strains with differences in their EPSs production. Strains recommended for inoculation as well as laboratory strains and native strains isolated from alfalfa fields were investigated. This study concentrated on EPS-II production, which results in mucoid colonies that are dependent on the presence of an intact expR gene. The results revealed that although the studied strains exhibited different phenotypes, the differences did not affect alfalfa nodulation itself. However, subtle changes in timing and efficacy to the effects of inoculation with the different strains may result because of other as-yet unknown factors. Thus, additional research is needed to determine the most effective inoculant strains and the best conditions for improving alfalfa production under agricultural conditions.