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Analysis of Rhizobia from root nodules

Rhizobia are soil bacteria capable of fixing atmospheric nitrogen when living in symbiosis with legumes inside root nodules. Nodulation is the fascinating result of complex interactions between the bacteria and the host.
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Plant-Microbe Symbioses
Symbiotic associations between plants and microbes
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E151 (sym15), a pleiotropic mutant of pea (Pisum sativum L.), displays low nodule number, enhanced mycorrhizae, delayed lateral root emergence, and high root cytokinin levels

E151 (sym15), a pleiotropic mutant of pea (Pisum sativum L.), displays low nodule number, enhanced mycorrhizae, delayed lateral root emergence, and high root cytokinin levels | Plant-Microbe Symbioses | Scoop.it
In legumes, the formation of rhizobial and mycorrhizal root symbioses is a highly regulated process which requires close communication between plant and microorganism. Plant mutants that have difficulties establishing symbioses are valuable tools for unravelling the mechanisms by which these symbioses are formed and regulated. Here E151, a mutant of Pisum sativum cv. Sparkle, was examined to characterize its root growth and symbiotic defects. The symbioses in terms of colonization intensity, functionality of micro-symbionts, and organ dominance were compared between the mutant and wild type. The endogenous cytokinin (CK) and abscisic acid (ABA) levels and the effect of the exogenous application of these two hormones were determined. E151 was found to be a low and delayed nodulator, exhibiting defects in both the epidermal and cortical programmes though a few mature and functional nodules develop. Mycorrhizal colonization of E151 was intensified, although the fungal functionality was impaired. Furthermore, E151 displayed an altered lateral root (LR) phenotype compared with that of the wild type whereby LR emergence is initially delayed but eventually overcome. No differences in ABA levels were found between the mutant and the wild type, but non-inoculated E151 exhibited significantly high CK levels. It is hypothesized that CK plays an essential role in differentially mediating the entry of the two micro-symbionts into the cortex; whereas it would inhibit the entry of the rhizobia in that tissue, it would promote that of the fungus. E151 is a developmental mutant which may prove to be a useful tool in further understanding the role of hormones in the regulation of beneficial root symbioses.
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Is there anything more beautiful the a nitrogen-fixing root nodule?... OK.. maybe an arbuscule :-)

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Metabolic dependencies drive species co-occurrence in diverse microbial communities

Metabolic dependencies drive species co-occurrence in diverse microbial communities | Plant-Microbe Symbioses | Scoop.it

Although metabolic interactions have long been implicated in the assembly of microbial communities, their general prevalence has remained largely unknown. In this study, we systematically survey, by using a metabolic modeling approach, the extent of resource competition and metabolic cross-feeding in over 800 microbial communities from diverse habitats. We show that interspecies metabolic exchanges are widespread in natural communities, and that such exchanges can provide group advantage under nutrient-poor conditions. Our results highlight metabolic dependencies as a major driver of species co-occurrence. The presented methodology and mechanistic insights have broad implications for understanding compositional variation in natural communities as well as for facilitating the design of synthetic microbial communities.


Via Kemen Lab, Stéphane Hacquard, Francis Martin
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Hsiao-Han Lin's curator insight, May 5, 9:51 PM

microbe are not the same, and they communicate and exchange

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Researchers sequence and analyze a cyanobacterium with known nitrogen-fixing capabilities

Researchers sequence and analyze a cyanobacterium with known nitrogen-fixing capabilities | Plant-Microbe Symbioses | Scoop.it
Researchers sequenced and analyzed the uncommonly large genome of a nitrogen-fixing cyanobacterial strain, comparing it to the more than 150 extant cyanobacterial genomes.
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Assessment of arbuscular mycorrhizal fungi status and heavy metal accumulation characteristics of tree species in a lead–zinc mine area: potential applications for phytoremediation

Assessment of arbuscular mycorrhizal fungi status and heavy metal accumulation characteristics of tree species in a lead–zinc mine area: potential applications for phytoremediation | Plant-Microbe Symbioses | Scoop.it
To select suitable tree species associated with arbuscular mycorrhizal fungi (AMF) for phytoremediation of heavy metal (HM) contaminated area, we measured the AMF status and heavy metal accumulation in plant tissues in a lead–zinc mine area, Northwest China. All 15 tree species were colonized by AM fungi in our investigation. The mycorrhizal frequency (F%), mycorrhizal colonization intensity (M%) and spore density (SP) reduced concomitantly with increasing Pb and Zn levels; however, positive correlations were found between arbuscule density (A%) and soil total/DTPA-extractable Pb concentrations. The average concentrations of Pb, Zn, Cu and Cd in plant samples were 168.21, 96.61, 41.06, and 0.79 mg/kg, respectively. Populus purdomii Rehd. accumulated the highest concentrations of Zn (432.08 mg/kg) and Cu (140.85 mg/kg) in its leaves. Considerable amount of Pb (712.37 mg/kg) and Cd (3.86 mg/kg) were concentrated in the roots of Robinia pseudoacacia Linn. and Populus simonii Carr., respectively. Plants developed different strategies to survive in HM stress environment: translocating more essential metals (Zn and Cu) into the aerial parts, while retaining more toxic heavy metals (Pb and Cd) in the roots to protect the above-ground parts from damage. According to the translocation factor (TF), bioconcentration factor (BCF), growth rate and biomass production, five tree species (Ailanthus altissima (Mill.) Swingle, Cotinus coggygria Scop., P. simonii, P. purdomii, and R. pseudoacacia) were considered to be the most suitable candidates for phytoextraction and/or phytostabilization purposes. Redundancy analysis (RDA) showed that the efficiency of phytoremediation was enhanced by AM symbioses, and soil pH, Pb, Zn, and Cd levels were the main factors influencing the HM accumulation characteristics of plants.
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Evolutionary tradeoffs as opportunities to improve yield potential

Photosynthetic efficiency and stress tolerance are examples of traits that had been improved by natural selection for millions of years prior to domestication of crops. Further improving such traits often requires accepting tradeoffs that would have reduced fitness of the crop's ancestors where they evolved. For example, improvements in yield potential have mostly come from reversing past selection for individual-plant competitiveness that conflicted with plant-community efficiency, or from tradeoffs between adaptation to past versus present conditions. A brief review of cold- and drought-tolerance did not find evidence of tradeoff-free improvements in crops, relative to wild ancestors. Identifying evolutionary tradeoffs that impose minimal agronomic tradeoffs can point the way to further improvements in yield potential and other community-level traits, perhaps including weed suppression. Crop genotypes that benefit subsequent crops merit more attention. Radical innovations never tested by natural selection may have considerable potential, but both tradeoffs and synergies will often be hard to predict.
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Influence of plant communities and soil properties during natural vegetation restoration on arbuscular mycorrhizal fungal communities in a karst region

Influence of plant communities and soil properties during natural vegetation restoration on arbuscular mycorrhizal fungal communities in a karst region | Plant-Microbe Symbioses | Scoop.it
Arbuscular mycorrhizal (AM) fungi are crucial for vegetation regeneration because they play an important role in plant establishment and growth. We investigated the relationships among AM fungal diversity, plant diversity, and soil properties during vegetation restoration in degraded karst ecosystems. Pyrosequencing approach was used to determine the genetic diversity and composition of AM fungal communities in four typical ecosystems of vegetation restoration (tussock (TK), shrub (SB), secondary forest (SF), and primary forest (PF)) in a karst region of southwest China. The diversity, richness, and evenness of plant species were evaluated through field surveys. Basic soil properties were measured. Plant species diversity and soil nutrient contents increased with vegetation restoration from tussock to primary forest, but the diversity of AM fungi followed the order of shrub ≈ secondary forest ≈ primary forest < tussock. The composition of AM fungi and plant communities differed significantly between ecosystems (p < 0.05). The richness of AM fungi was negatively correlated with both the plant diversity (the indices of plant Shannon–Wiener, evenness and richness) and soil properties (soil available phosphorus (AP), soil organic carbon (SOC) and pH) (p < 0.05). Redundancy analysis showed that the AM fungal communities closely linked to plant richness, soil organic carbon, soil available phosphorus and pH. These results suggest that the diversity and composition of AM fungi in karst region are influenced by plant communities and soil nutrient conditions.
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Agricultural microbiology draws Chinese researchers to Argentina

Agricultural microbiology draws Chinese researchers to Argentina | Plant-Microbe Symbioses | Scoop.it
Microbiology applied to agricultural products is connecting China with Argentina, two emerging markets with complementary economies and strategic partners looking to increase bilateral exchanges.

Among the many links between Beijing and Buenos Aires, Argentine firm Rizobacter stands out. The company, which uses microbiology to boost soy output while cutting production costs, has been in business for 38 years and reports an annual turnover of US$100 million, 20% from exports.

In December, a delegation of the Academy of Sciences from China's Heilongjiang province visited the firm in Pergamino, a city located 180 km northeast of the capital Buenos Aires.

The delegation came to explore the possibility of signing an agreement for joint research and development of microbiological technologies, to improve Chinese soil and output.

"We are here to visit and get to know the Rizobacter plant," said Wang Gang, vice president of the academy.

"We are very interested in getting to know the technology being developed by this company and all of the efforts on the production of soybean, mainly related to soybeans and rhizobia," Wang added.
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Role of mycorrhization and nutrient availability in competitive interactions between the grassland species Plantago lanceolata and Hieracium pilosella

Role of mycorrhization and nutrient availability in competitive interactions between the grassland species Plantago lanceolata and Hieracium pilosella | Plant-Microbe Symbioses | Scoop.it
Arbuscular mycorrhizal fungi (AMF) may serve as an effective substitute for root surface. As mycorrhizal benefits are related to nutrient availability, the trade-off between carbon investments into AMF versus roots may drive competitive interactions. We studied competitive interactions between mycorrhizal and non-mycorrhizal individuals of Hieracium pilosella L. and Plantago lanceolata L., species differing in both mycotrophic degree and carbon allocation to roots. Three fertilization treatments were used to simulate nutritional differences over the course of succession. Species-specific differences in mycotrophy were reflected in markedly larger root/shoot allocation in P. lanceolata and higher mycorrhizal growth dependency in H. pilosella. P. lanceolata dominated competition in all fertilizer treatments, enabled by its comparatively larger root biomass allocation. In contrast, under intermediate and high fertilization, H. pilosella exhibited large investments into clonal shoot growth rather than in roots. Unexpectedly, the competitive imbalance between both species was amplified by the presence of AMF. The poor competitive strength of H. pilosella indicates that AMF-dominated foraging can be less effective than root-dominated foraging in competitive interactions, particularly under high nutrient availabilities. However, the competitive imbalance was reduced in favor of H. pilosella under nutrient deficiency. Our results lend support to the idea of differing competitive success of mycorrhizal- versus root-based foraging strategy over a nutritional gradient, which may play a role in the natural distribution of species over the course of succession.
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Multiple Autoregulation of Nodulation (AON) Signals Identified through Split Root Analysis of Medicago truncatula sunn and rdn1 Mutants

Multiple Autoregulation of Nodulation (AON) Signals Identified through Split Root Analysis of Medicago truncatula sunn and rdn1 Mutants | Plant-Microbe Symbioses | Scoop.it
Nodulation is energetically costly to the host: legumes balance the nitrogen demand with the energy expense by limiting the number of nodules through long-distance signaling. A split root system was used to investigate systemic autoregulation of nodulation (AON) in Medicago truncatula and the role of the AON genes RDN1 and SUNN in the regulatory circuit. Developing nodule primordia did not trigger AON in plants carrying mutations in RDN1 and SUNN genes, while wild type plants had fully induced AON within three days. However, despite lacking an early suppression response, AON mutants suppressed nodulation when roots were inoculated 10 days or more apart, correlated with the maturation of nitrogen fixing nodules. In addition to correlation between nitrogen fixation and suppression of nodulation, suppression by extreme nutrient stress was also observed in all genotypes and may be a component of the observed response due to the conditions of the assay. These results suggest there is more than one systemic regulatory circuit controlling nodulation in M. truncatula. While both signals are present in wild type plants, the second signal can only be observed in plants lacking the early repression (AON mutants). RDN1 and SUNN are not essential for response to the later signal.
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EcoBeneficial Interview with Dr. Roger Koide on Mycorrhizal Fungi

Join Kim Eierman, Founder of EcoBeneficial, for an interview with Dr. Roger Koide, professor at Brigham Young University for a discussion about mycorrhizal ...
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Molecular principles of membrane microdomain targeting in plants

Molecular principles of membrane microdomain targeting in plants | Plant-Microbe Symbioses | Scoop.it
Plasma membranes (PMs) are heterogeneous lipid bilayers comprising diverse subdomains. These sites can be labeled by various proteins in vivo and may serve as hotspots for signal transduction. They are found at apical, basal, and lateral membranes of polarized cells, at cell equatorial planes, or almost isotropically distributed throughout the PM. Recent advances in imaging technologies and understanding of mechanisms that allow proteins to target specific sites in PMs have provided insights into the dynamics and complexity of their specific segregation. Here we present a comprehensive overview of the different types of membrane microdomain and describe the molecular modes that determine site-directed targeting of membrane-resident proteins at the PM.
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Very nice review

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Mapping Genomes of Pine Bacteria Yields Unexpected Results

Mapping Genomes of Pine Bacteria Yields Unexpected Results | Plant-Microbe Symbioses | Scoop.it
UC Merced Professor Carolin Frank is helping figure out how a certain bacteria helps promote healthy tree growth by studying the bacteria’s genome sequence.

In the new paper, published in the open-access journal mBioOpens a new window, Frank and research colleagues in Finland said their discoveries could have practical applications in agriculture and forestry by stimulating plant growth.

While trying to grow pine tree cells in petri dishes, the Finnish researchers couldn’t get rid of the Methylobacterium extorquens microbe. They were surprised to find the bacterium collecting inside the tree’s cells, especially around the nuclei.

And they found that inoculating the pine seedlings with the bacteria made the little trees grow faster, suggesting the bacteria are beneficial. That made Frank want to unveil the mechanisms behind the host-microbe interaction.
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microRNA167-directed regulation of the auxin response factors, GmARF8a and GmARF8b, is required for soybean (Glycine max L.) nodulation and lateral root development

Legumes root nodules convert atmospheric nitrogen gas (N2) into ammonium through symbiosis with a prokaryotic microsymbiont broadly called 'rhizobia'. Auxin signaling is required for determinant nodule development; however, the molecular mechanism of auxin-mediated nodule formation remains largely unknown. Here we show in soybean (Glycine max L. Merr) that microRNA (miRNA) miR167 acts as a positive regulator of lateral root organs, namely nodules and lateral roots. miR167c expression was up-regulated in the vasculature, pericycle and cortex of soybean roots following inoculation with Bradyrhizobium japonicum strain USDA110 (the microsymbiont). It was found to positively regulate nodule numbers directly by repressing the target genes GmARF8a and GmARF8b (homologous genes of Arabidopsis AtARF8 that encode auxin response factors). Moreover, the expression of miR167 and its targets were up- or down-regulated by auxin, respectively. The miR167-GmARF8 module also positively regulated nodulation efficiency under low microsymbiont density, a condition often associated with environmental stress. The regulatory role of miR167 on nodule initiation was dependent on the Nod factor receptor GmNFR1α, and it acts upstream of the nodulation-associated genes GmNIN, GmNSP1, GmENOD40-1, GmHAP2-1 and GmHAP2-2. miR167 also promoted lateral root numbers. Collectively, our findings establish a key role for the miR167-GmARF8 module in auxin-mediated nodule and lateral root formation in soybean.
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The private life of a common garden weed

The private life of a common garden weed | Plant-Microbe Symbioses | Scoop.it
The umbrella liverwort (Marchantia polymorpha) is a noxious weed. It grows in most gardens, paths and glasshouses, and is found in almost all parts of the world – from tropical to arctic climates. There are several subspecies of this plant that are found in damp habitats at varying altitudes. The plants themselves are of little economic importance, but in ancient times, liverworts were believed to be a cure for liver diseases. This ancient idea was part of the “doctrine of signatures” [1]. This doctrine suggested that plants resembling a certain body part could be used to treat ailments of that body part. Although an interesting idea, there is no scientific evidence to support this claim.
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Math, models, microbes: Diving into the complex soil community interactions of plant-soil feedbacks

Math, models, microbes: Diving into the complex soil community interactions of plant-soil feedbacks | Plant-Microbe Symbioses | Scoop.it
A major challenge for predicting the strength and direction of plant-soil feedbacks (PSF) is understanding the interactions between plant traits, belowground communities, and the soil processes both compartments influence. A new article by Ke et al. (2015) uses a modeling approach to test how interactions between litter- and root-mediated pathways can influence which plant and soil biota traits are important mechanisms that drive PSF. Here, we will unpack their model in a simple way and touch on some of the interesting ecological and evolutionary implications.
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Phosphoproteomics in Cereals

Phosphoproteomics in Cereals | Plant-Microbe Symbioses | Scoop.it
Cereals are the most important crop plant supplying staple food throughout the world. The economic importance and continued breeding of crop plants such as rice, maize, wheat, or barley require a detailed scientific understanding of adaptive and developmental processes. Protein phosphorylation is one of the most important regulatory posttranslational modifications and its analysis allows deriving functional and regulatory principles in plants. This minireview summarizes the current knowledge of phosphoproteomic studies in cereals.
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Evaluating the use of plant hormones and biostimulators in forage pastures to enhance shoot dry biomass production by perennial ryegrass (Lolium perenne L.)

Fertilization of established perennial ryegrass forage pastures with nitrogen (N) - based fertilizers is currently the most common practice used on farms to increase pasture forage biomass yield. However, over-fertilization can lead to undesired environmental impacts, including nitrate leaching into waterways and increased gaseous emissions of ammonia and nitrous oxide to the atmosphere. Additionally, there is growing interest from pastoral farmers to adopt methods for increasing pasture dry matter yield which use ‘natural’, environmentally safe plant growth stimulators, together with N-based fertilizers. Such plant growth stimulators include plant hormones and plant growth promotive micro-organisms such as bacteria and fungi (‘biostimulators’, which may produce plant growth-inducing hormones), as well as extracts of seaweed (marine algae). This review presents examples and discusses current uses of plant hormones and biostimulators, applied alone or together with N-based fertilizers, to enhance shoot dry matter yield of forage pasture species, with an emphasis on perennial ryegrass.
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Researchers share the dirt on Year of Soils

Researchers share the dirt on Year of Soils | Plant-Microbe Symbioses | Scoop.it
We walk on it, we eat from it, we draw our water through it, and we send our waste into its depths to return clean once more.

But what do we really know about soil?
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The carbon starvation response of the ectomycorrhizal fungus Paxillus involutus

The carbon starvation response of the ectomycorrhizal fungus Paxillus involutus | Plant-Microbe Symbioses | Scoop.it
The amounts of carbon allocated to the fungal partner in ectomycorrhizal associations can vary substantially depending on the plant growth and the soil nutrient conditions, and the fungus may frequently be confronted with limitations in carbon. We used chemical analysis and transcriptome profiling to examine the physiological response of the ectomycorrhizal fungus Paxillus involutus to carbon starvation during axenic cultivation. Carbon starvation induced a decrease in the biomass. Concomitantly, ammonium, cell wall material (chitin) and proteolytic enzymes were released into the medium, which suggest autolysis. Compared with the transcriptome of actively growing hyphae, about 45% of the transcripts analyzed were differentially regulated during C-starvation. Induced during starvation were transcripts encoding extracellular enzymes such as peptidases, chitinases and laccases. In parallel, transcripts of N-transporters were upregulated, which suggest that some of the released nitrogen compounds were re-assimilated by the mycelium. The observed changes suggest that the carbon starvation response in P. involutus is associated with complex cellular changes that involves autolysis, recycling of intracellular compounds by autophagy and reabsorption of the extracellular released material. The study provides molecular markers that can be used to examine the role of autolysis for the turnover and survival of the ectomycorrhizal mycelium in soils.
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Mutualistic root endophytism is not associated with the reduction of saprotrophic traits and requires a noncompromised plant innate immunity

During a compatible interaction, the sebacinoid root-associated fungi Piriformospora indica and Sebacina vermifera induce modification of root morphology and enhance shoot growth in Arabidopsis thaliana.
The genomic traits common in these two fungi were investigated and compared with those of other root-associated fungi and saprotrophs. The transcriptional responses of the two sebacinoid fungi and of Arabidopsis roots to colonization at three different symbiotic stages were analyzed by custom-designed microarrays.
We identified key genomic features characteristic of sebacinoid fungi, such as expansions for gene families involved in hydrolytic activities, carbohydrate-binding and protein–protein interaction. Additionally, we show that colonization of Arabidopsis correlates with the induction of salicylic acid catabolism and accumulation of jasmonate and glucosinolates (GSLs). Genes involved in root developmental processes were specifically induced by S. vermifera at later stages during interaction.
Using different Arabidopsis indole-GSLs mutants and measurement of secondary metabolites, we demonstrate the importance of the indolic glucosinolate pathway in the growth restriction of P. indica and S. vermifera and we identify indole-phytoalexins and specifically indole-carboxylic acids derivatives as potential key players in the maintenance of a mutualistic interaction with root endophytes.
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Molecular adaptations of Herbaspirillum seropedicae during colonization of the maize rhizosphere

Molecular adaptations of Herbaspirillum seropedicae during colonization of the maize rhizosphere | Plant-Microbe Symbioses | Scoop.it
Molecular mechanisms of plant recognition and colonization by diazotrophic bacteria are barely understood. Herbaspirillum seropedicae is a Betaproteobacterium capable of colonizing epiphytically and endophytically commercial grasses, to promote plant growth. In this study, we utilized RNA-seq to compare the transcriptional profiles of planktonic and maize root-attached H. seropedicae SmR1 recovered 1 and 3 days after inoculation. The results indicated that nitrogen metabolism was strongly activated in the rhizosphere and polyhydroxybutyrate storage was mobilized in order to assist survival of H. seropedicae during the early stages of colonization. Epiphytic cells showed altered transcription levels of several genes associated with polysaccharide biosynthesis, peptidoglycan turnover and outer membrane protein biosynthesis, suggesting reorganization of cell wall envelope components. Specific methyl-accepting chemotaxis proteins and two-component systems were differentially expressed between populations over time, suggesting deployment of an extensive bacterial sensory system for adaptation to the plant environment. An insertion mutation inactivating a methyl-accepting chemosensor induced in planktonic bacteria, decreased chemotaxis towards the plant and attachment to roots. In summary, analysis of mutant strains combined with transcript profiling revealed several molecular adaptations that enable H. seropedicae to sense the plant environment, attach to the root surface and survive during the early stages of maize colonization.
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Silencing a key gene of the common symbiosis pathway in Nicotiana attenuata specifically impairs arbuscular mycorrhizal infection without influencing the root-associated microbiome or plant growth.

Silencing a key gene of the common symbiosis pathway in Nicotiana attenuata specifically impairs arbuscular mycorrhizal infection without influencing the root-associated microbiome or plant growth. | Plant-Microbe Symbioses | Scoop.it
While the biochemical function of calcium and calmodulin-dependent protein kinase (CCaMK) is well studied, and plants impaired in the expression of CCaMK are known not to be infected by arbuscular mycorrhizal fungi (AMF) in glasshouse studies, the whole-plant and ecological consequences of CCaMK silencing are not well understood. Here we show that three independently transformed lines of Nicotiana attenuata plants silenced in CCaMK (irCCaMK) are neither infected by Rhizophagus irregularis in the glasshouse nor by native fungal inoculum in the field. The overall fungal community of field-grown roots did not differ significantly among empty vector (EV) and the transgenic lines, and the bacterial communities only showed minor differences, as revealed by the alpha diversity parameters of bacterial OTUs, which were higher in EV plants compared to two of the three transformed lines, while beta-diversity parameters did not differ. Furthermore, growth and fitness parameters were similar in the glasshouse and field. Herbivory-inducible and basal levels of salicylic acid, jasmonic acid and abscisic acid did not differ amongst the genotypes, suggesting that activation of the classical defense pathways are not affected by CCaMK silencing. Based on these results we conclude that silencing of CCaMK has few, if any, non-target effects.

Via Francis Martin, Christophe Jacquet
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That's very interesting.

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Pedobiologia: Journal of Soil Ecology's curator insight, April 30, 7:20 PM

A useful model for disentagling the interactive effects of AM fungi and other microbes on plant community dynamics and trophic interactions?

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The Intracellular Scots Pine Shoot Symbiont Methylobacterium extorquens DSM13060 Aggregates around the Host Nucleus and Encodes Eukaryote-Like Proteins

The Intracellular Scots Pine Shoot Symbiont Methylobacterium extorquens DSM13060 Aggregates around the Host Nucleus and Encodes Eukaryote-Like Proteins | Plant-Microbe Symbioses | Scoop.it
Endophytes are microbes that inhabit plant tissues without any apparent signs of infection, often fundamentally altering plant phenotypes. While endophytes are typically studied in plant roots, where they colonize the apoplast or dead cells, Methylobacterium extorquens strain DSM13060 is a facultatively intracellular symbiont of the meristematic cells of Scots pine (Pinus sylvestris L.) shoot tips. The bacterium promotes host growth and development without the production of known plant growth-stimulating factors. Our objective was to examine intracellular colonization by M. extorquens DSM13060 of Scots pine and sequence its genome to identify novel molecular mechanisms potentially involved in intracellular colonization and plant growth promotion. Reporter construct analysis of known growth promotion genes demonstrated that these were only weakly active inside the plant or not expressed at all. We found that bacterial cells accumulate near the nucleus in intact, living pine cells, pointing to host nuclear processes as the target of the symbiont’s activity. Genome analysis identified a set of eukaryote-like functions that are common as effectors in intracellular bacterial pathogens, supporting the notion of intracellular bacterial activity. These include ankyrin repeats, transcription factors, and host-defense silencing functions and may be secreted by a recently imported type IV secretion system. Potential factors involved in host growth include three copies of phospholipase A2, an enzyme that is rare in bacteria but implicated in a range of plant cellular processes, and proteins putatively involved in gibberellin biosynthesis. Our results describe a novel endophytic niche and create a foundation for postgenomic studies of a symbiosis with potential applications in forestry and agriculture.

IMPORTANCE All multicellular eukaryotes host communities of essential microbes, but most of these interactions are still poorly understood. In plants, bacterial endophytes are found inside all tissues. M. extorquens DSM13060 occupies an unusual niche inside cells of the dividing shoot tissues of a pine and stimulates seedling growth without producing cytokinin, auxin, or other plant hormones commonly synthesized by plant-associated bacteria. Here, we tracked the bacteria using a fluorescent tag and confocal laser scanning microscopy and found that they localize near the nucleus of the plant cell. This prompted us to sequence the genome and identify proteins that may affect host growth by targeting processes in the host cytoplasm and nucleus. We found many novel genes whose products may modulate plant processes from within the plant cell. Our results open up new avenues to better understand how bacteria assist in plant growth, with broad implications for plant science, forestry, and agriculture.
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That's unusual to say the least!

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