Project description:Rhizosphere is a complex system of interactions between plant roots, bacteria, fungi and animals, where the release of plant root exudates stimulates bacterial density and diversity. However, the majority of the bacteria in soil results to be unculturable but active. The aim of the present work was to characterize the microbial community associated to the root of V. vinifera cv. Pinot Noir not only under a taxonomic perspective, but also under a functional point of view, using a metaproteome approach. Our results underlined the difference between the metagenomic and metaproteomic approach and the large potentiality of proteomics in describing the environmental bacterial community and its activity. In fact, by this approach, that allows to investigate the mechanisms occurring in the rhizosphere, we showed that bacteria belonging to Streptomyces, Bacillus and Pseudomonas genera are the most active in protein expression. In the rhizosphere, the identified genera were involved mainly in phosphorus and nitrogen soil metabolism.
Project description:Plants and rhizosphere microbes rely closely on each other, with plants supplying carbon to bacteria in root exudates, and bacteria mobilizing soil-bound phosphate for plant nutrition. When the phosphate supply becomes limiting for plant growth, the composition of root exudation changes, affecting rhizosphere microbial communities and microbially-mediated nutrient fluxes. To evaluate how plant phosphate deprivation affects rhizosphere bacteria, Lolium perenne seedlings were root-inoculated with Pseudomonas aeruginosa 7NR, and grown in axenic microcosms under different phosphate regimes (330 uM vs 3-6 uM phosphate). The effect of biological nutrient limitation was examined by DNA microarray studies of rhizobacterial gene expression.
Project description:Nipponbare performs poorly in phosphorus (P) deficient soil whereas a Nipponbare-derived NIL containing the Pup1 allele of donor parent Kasalath is tolerant to P deficiency. In this experiment we compared gene expression patterns in roots of this NIL to Nipponbare, grown either in a P deficient or P fertilized soil. The aim is to separate constitutive differences in expression from those induced by P deficiency. Keywords: genotype comparison, constitutive differential expression
Project description:Advances in DNA sequencing technologies has drastically changed our perception of the structure and complexity of the plant microbiome. By comparison, our ability to accurately identify the metabolically active fraction of soil microbiota and its specific functional role in augmenting plant health is relatively limited. Here, we combined our recently developed protein extraction method and an iterative bioinformatics pipeline to enable the capture and identification of extracellular proteins (metaexoproteomics) synthesised in the rhizosphere of Brassica spp. We first validated our method in the laboratory by successfully identifying proteins related to a host plant (Brassica rapa) and its bacterial inoculant, Pseudomonas putida BIRD-1. This identified numerous rhizosphere specific proteins linked to the acquisition of plant-derived nutrients in P. putida. Next, we analysed natural field-soil microbial communities associated with Brassica napus L. (oilseed rape). By combining metagenomics with metaexoproteomics, 1882 proteins were identified across bulk and rhizosphere samples. Meta-exoproteomics identified a clear shift (p<0.001) in the metabolically active fraction of the soil microbiota responding to the presence of B. napus roots that was not apparent in the composition of the total microbial community (metagenome). This metabolic shift was associated with the stimulation of rhizosphere-specialised bacteria, such as Gammaproteobacteria, Betaproteobacteria and Flavobacteriia and the upregulation of plant beneficial functions related to phosphorus and nitrogen mineralisation. Together, our metaproteomic assessment of the ‘active’ plant microbiome at the field-scale demonstrates the importance of moving past a genomic assessment of the plant microbiome in order to determine ecologically important plant-microbe interactions underpinning plant health.
Project description:Advances in DNA sequencing technologies has drastically changed our perception of the structure and complexity of the plant microbiome. By comparison, our ability to accurately identify the metabolically active fraction of soil microbiota and its specific functional role in augmenting plant health is relatively limited. Here, we combined our recently developed protein extraction method and an iterative bioinformatics pipeline to enable the capture and identification of extracellular proteins (metaexoproteomics) synthesised in the rhizosphere of Brassica spp. We first validated our method in the laboratory by successfully identifying proteins related to a host plant (Brassica rapa) and its bacterial inoculant, Pseudomonas putida BIRD-1. This identified numerous rhizosphere specific proteins linked to the acquisition of plant-derived nutrients in P. putida. Next, we analysed natural field-soil microbial communities associated with Brassica napus L. (oilseed rape). By combining metagenomics with metaexoproteomics, 1882 proteins were identified across bulk and rhizosphere samples. Meta-exoproteomics identified a clear shift (p<0.001) in the metabolically active fraction of the soil microbiota responding to the presence of B. napus roots that was not apparent in the composition of the total microbial community (metagenome). This metabolic shift was associated with the stimulation of rhizosphere-specialised bacteria, such as Gammaproteobacteria, Betaproteobacteria and Flavobacteriia and the upregulation of plant beneficial functions related to phosphorus and nitrogen mineralisation. Together, our metaproteomic assessment of the ‘active’ plant microbiome at the field-scale demonstrates the importance of moving past a genomic assessment of the plant microbiome in order to determine ecologically important plant-microbe interactions underpinning plant health.
Project description:Advances in DNA sequencing technologies has drastically changed our perception of the structure and complexity of the plant microbiome. By comparison, our ability to accurately identify the metabolically active fraction of soil microbiota and its specific functional role in augmenting plant health is relatively limited. Here, we combined our recently developed protein extraction method and an iterative bioinformatics pipeline to enable the capture and identification of extracellular proteins (metaexoproteomics) synthesised in the rhizosphere of Brassica spp. We first validated our method in the laboratory by successfully identifying proteins related to a host plant (Brassica rapa) and its bacterial inoculant, Pseudomonas putida BIRD-1. This identified numerous rhizosphere specific proteins linked to the acquisition of plant-derived nutrients in P. putida. Next, we analysed natural field-soil microbial communities associated with Brassica napus L. (oilseed rape). By combining metagenomics with metaexoproteomics, 1882 proteins were identified across bulk and rhizosphere samples. Meta-exoproteomics identified a clear shift (p<0.001) in the metabolically active fraction of the soil microbiota responding to the presence of B. napus roots that was not apparent in the composition of the total microbial community (metagenome). This metabolic shift was associated with the stimulation of rhizosphere-specialised bacteria, such as Gammaproteobacteria, Betaproteobacteria and Flavobacteriia and the upregulation of plant beneficial functions related to phosphorus and nitrogen mineralisation. Together, our metaproteomic assessment of the ‘active’ plant microbiome at the field-scale demonstrates the importance of moving past a genomic assessment of the plant microbiome in order to determine ecologically important plant-microbe interactions underpinning plant health.
Project description:Aluminum (Al)–tolerant phosphobacteria can improve plant performance in acidic soils by increasing Al complexing and phosphorus (P) availability. However, it is almost unknown how Al stress along with P deficiency affect the bacterial biochemistry and physiology. Because high Al levels and low P availability often occur simultaneously in acidic soils, we have evaluated the single and mutual effects of a high Al stress and P deficiency on the proteome of the Al‒tolerant phosphobacteria strain Enterobacter sp. RJAL6. This strain was previously isolated from the rhizosphere of Lollium perenne plants grown in acidic soil. The strain was cultivated in mineral media modified to contain i) high P (1.4 mM) in the absence of Al, ii) high P (1.4 mM) and high Al (10 mM), iii) low P (0.05 mM) in the absence of Al, and iv) low P (0.05 mM) and high Al (10 mM). Total proteins from bacterial cells were extracted at the end of the exponential phase of growth and subjected to high–throughput proteomics analysis. The results showed that P deficiency was mainly associated with an upregulation of the P metabolism proteins subject to Pho regulon control, including phosphatases and transporters involved in the uptake of organophosphorus compounds such as phosphomonoesters, phosphonates and glycerol–3–phosphate. Aluminum exposure primarily decreased the expression of iron (Fe)–sulfur and haem-containing proteins with a concomitant upregulation of Fe acquisition and metabolism proteins, including siderophore precursors and receptors of Fe–chelator complexes. Here, we demonstrated the preponderant role that Al plays in the adjustment of Fe homeostasis, and consequently in the central metabolism of the bacteria. This is the first report of a proteomic study of the interaction between high Al and P deficiency in acidic soil–adapted bacteria. This knowledge is crucial for developing bioinoculants for crops affected by both Al toxicity and P deficiency.
Project description:Aluminum (Al)–tolerant phosphobacteria can improve plant performance in acidic soils by increasing Al complexing and phosphorus (P) availability. However, it is almost unknown how Al stress along with P deficiency affect the bacterial biochemistry and physiology. Because high Al levels and low P availability often occur simultaneously in acidic soils, we have evaluated the single and mutual effects of a high Al stress and P deficiency on the proteome of the Al‒tolerant phosphobacteria strain Enterobacter sp. 198. This strain was previously isolated from the rhizosphere of Lollium perenne plants grown in acidic soil. The strain was cultivated in mineral media modified to contain i) high P (1.4 mM) in the absence of Al, ii) high P (1.4 mM) and high Al (10 mM), iii) low P (0.05 mM) in the absence of Al, and iv) low P (0.05 mM) and high Al (10 mM). Total proteins from bacterial cells were extracted at the end of the exponential phase of growth and subjected to high–throughput proteomics analysis. The results showed that P deficiency was mainly associated with an upregulation of the P metabolism proteins subject to Pho regulon control, including phosphatases and transporters involved in the uptake of organophosphorus compounds such as phosphomonoesters, phosphonates and glycerol–3–phosphate. Aluminum exposure primarily decreased the expression of iron (Fe)–sulfur and haem-containing proteins with a concomitant upregulation of Fe acquisition and metabolism proteins, including siderophore precursors and receptors of Fe–chelator complexes. Here, we demonstrated the preponderant role that Al plays in the adjustment of Fe homeostasis, and consequently in the central metabolism of the bacteria. This is the first report of a proteomic study of the interaction between high Al and P deficiency in acidic soil–adapted bacteria. This knowledge is crucial for developing bioinoculants for crops affected by both Al toxicity and P deficiency.