Interactions between Pseudomonas putida UW4 and Gigaspora rosea BEG9 and their consequences for the growth of cucumber under salt-stress conditions

Aims: After the determination of the toxic but nonlethal concentration of NaCl for cucumber, we examined the interaction between an ACC (1‐aminocyclopropane‐1‐carboxylate) deaminase producing bacterial strain and an arbuscular mycorrhizal fungus (AMF) and their effects on cucumber growth under salinity. Methods and Results: In the first experiment, cucumber seedlings were exposed to 0·1, 50, 100 or 200 mmol l^?1 NaCl, and plant biomass and leaf area were measured. While seeds exposed to 200 mmol l^?1 NaCl did not germinate, plant growth and leaf size were reduced by 50 or 100 mmol l^?1 salt. The latter salt cancentration caused plant death in 1 month. In the second experiment, seeds were inoculated with the ACC deaminase‐producing strain Pseudomonas putida UW4 (AcdS⁺), its mutant unable to produce the enzyme (AcdS^?), or the AMF Gigaspora rosea BEG9, individually or in combination and exposed to 75 mmol l^?1 salt. Plant morphometric and root architectural parameters, mycorrhizal and bacterial colonization and the influence of each micro‐organism on the photosynthetic efficiency were evaluated. The AcdS⁺ strain or the AMF, inoculated alone, increased plant growth, affected root architecture and improved photosynthetic activity. Mycorrhizal colonization was inhibited by each bacterial strain. Conclusions: Salinity negatively affects cucumber growth and health, but root colonization by ACC deaminase‐producing bacteria or arbuscular mycorrhizal fungi can improve plant tolerance to such stressful condition. Significance and Impact of the Study: Arbuscular mycorrhizal fungus and bacterial ACC deaminase may ameliorate plant growth under stressful conditions. It was previously shown that, under optimal growth conditions, Ps. putida UW4 AcdS⁺ increases root colonization by Gi. rosea resulting in synergistic effects on cucumber growth. These results suggest that while in optimal conditions ACC deaminase is mainly involved in the bacteria/fungus interactions, while under stressful conditions this enzyme plays a role in plant/bacterium interactions. This finding is relevant from an ecological and an applicative point of view.     

Bacteria with ACC deaminase can promote plant growth and help to feed the world

To feed all of the world's people, it is necessary to sustainably increase agricultural productivity. One way to do this is through the increased use of plant growth-promoting bacteria; recently, scientists have developed a more profound understanding of the mechanisms employed by these bacteria to facilitate plant growth. Here, it is argued that the ability of plant growth-promoting bacteria that produce 1-aminocyclopropane-1-carboxylate (ACC) deaminase to lower plant ethylene levels, often a result of various stresses, is a key component in the efficacious functioning of these bacteria. The optimal functioning of these bacteria includes the synergistic interaction between ACC deaminase and both plant and bacterial auxin, indole-3-acetic acid (IAA). These bacteria not only directly promote plant growth, they also protect plants against flooding, drought, salt, flower wilting, metals, organic contaminants, and both bacterial and fungal pathogens. While a considerable amount of both basic and applied work remains to be done before ACC deaminase-producing plant growth-promoting bacteria become a mainstay of plant agriculture, the evidence indicates that with the expected shift from chemicals to soil bacteria, the world is on the verge of a major paradigm shift in plant agriculture.     

Isolation and characterization of an unusual 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase gene from Enterobacter cloacae UW4

A genomic library of the 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase-containing plant growth-promoting bacterium Enterobacter cloacae UW4 in pUC19 in Escherichia coli was screened for the ability to utilize ACC as a sole source of nitrogen. One of the clones that was isolated contained a plasmid with an insert of approximately 0.8 kb that conferred ACC deaminase activity. Sequence analysis revealed that this DNA fragment contains an open-reading frame of 696 nucleotides predicted to encode a protein of 232 amino acids, a member of the amidohydrolase protein superfamily, i.e., a deaminase that contains a mononuclear or binuclear metal center as compared to the canonical ACC deaminase which contains pyridoxal phosphate as a co-factor.     

Mycorrhizal fungi influence plant and soil functions and interactions

Potted soybean (Glycine max (L.) Merr.) plants were grown in P-fertilized (+P) or low-P soil (-P), or colonized in -P soil by one of the arbuscular mycorrhizal (AM) fungi Glomus etunicatum (Ge), Glomus mosseae (Gm), or Gigaspora rosea (Gr). Treatment effects on plant development, on the soil microflora, and on the status of water-stable soil aggregates (WSA) were evaluated for all 5 treatments or for the 3 AM treatments alone. Dry weights of the AM plants, as a group, were half-way between the dry weights of the +P and -P plants, but within the AM group, Gm plants had the highest pod dry weights and pod/stem and root/stem ratios and the lowest specific root lengths, while Ge plants had high stem dry weights and were highly nodulated. High reproductive development and coarse roots in the Gm plants were associated with the most extensive growth of AM soil hyphae (km pot-1: Gm, 20; Gr, 12; Ge, 8), while nodulation was inversely related with AM-colonized root length. The soils colonized by AM fungi had significantly higher levels of WSA (size classes 1 to 2 and 2 to 4 mm), and within the larger size class, Gm soils had the highest percentage of WSA. Proliferation (plate counts) of Gram positive (G+) and Gram negative (G-) bacteria, Arthrobacter sp. (G+), and Pseudomonas sp. (G-) was greatest in the -P soils, but the bacterial populations of the +P and the AM soils were generally not significantly different. There were, however, differences among the AM treatments, where Gm soils had the lowest G- bacterial populations, while Ge soils had the highest populations of both G+ and G- bacteria. Correlations between plant and soil traits indicated that interactions within the plant-soil system were mediated by the AM fungi.     

The bacterium Paenibacillus validus stimulates growth of the arbuscular mycorrhizal fungus Glomus intraradices up to the formation of fertile spores

Two isolates of Paenibacillus validus (DSM ID617 and ID618) stimulated growth of the arbuscular mycorrhizal fungus Glomus intraradices Sy167 up to the formation of fertile spores, which recolonize carrot roots. Thus, the fungus was capable of completing its life cycle in the absence of plant roots, but relied instead on the simultaneous growth of bacteria. The supernatant of a mixed batch culture of the two P. validus isolates contained raffinose and another, unidentified trisaccharide. Among the oligosaccharides tested, raffinose was most effective in stimulating hyphal mass formation on plates but could not promote growth to produce fertile spores. A suppressive subtractive hybridization library followed by reverse Northern analyses indicated that several genes with products involved in signal transduction are differentially expressed in G. intraradices SY 167 when grown in coculture with P. validus (DSM 3037). The present investigation, while likely representing a significant step forward in understanding the arbuscular mycorrhizal fungus symbioses, also confirms that its optimal establishing and functioning might rely on many, as yet unidentified factors.     

Expression and characterization of 1-aminocyclopropane-1-carboxylate deaminase from the rhizobacterium Pseudomonas putida UW4: a key enzyme in bacterial plant growth promotion

The enzyme 1-aminocyclopropane-1-carboxylate deaminase (ACCD) converts ACC, the precursor of the plant hormone ethylene, to alpha-ketobutyrate and ammonium. This enzyme has been identified in soil bacteria and has been proposed to play a key role in microbe-plant association. A soluble recombinant ACCD from Pseudomonas putida UW4 of molecular weight 41 kDa has been cloned, expressed, and purified. It showed selectivity and high activity towards the substrate ACC: K(M)=3.4+/-0.2 mM and k(cat)=146+/-5 min(-1) at pH 8.0 and 22 degrees C. The enzyme displayed optimal activity at pH 8.0 with a sharp decline to essentially no activity below pH 6.5 and a slightly less severe tapering in activity at higher pH resulting in loss of activity at pH>10. The major component of the enzyme's secondary structure was determined to be alpha-helical by circular dichroism (CD). P. putida UW4 ACCD unfolded at 60 degrees C as determined by its CD temperature profile as well as by differential scanning microcalorimetry (DSC). Enzyme activity was knocked out in the point mutant Gly44Asp. Modeling this mutation into the known yeast ACCD structure shed light on the role this highly conserved residue plays in allowing substrate accessibility to the active site. This enzyme's biochemical and biophysical properties will serve as an important reference point to which newly isolated ACC deaminases from other organisms can be compared.     

The interface between the cell cycle and plant growth regulators: a mini review

The aim was to review knowledge about the interface betweenplant growth regulators and molecular checkpoints of the cell cycle. Atwhat level of biochemical regulation of the cell cycle do plant growthregulators interface? Are there different levels of interfacingdependent on the plant growth regulator involved? As a preamble totackling these questions, we overview the eukaryotic cell cycle withparticular emphasis on checkpoints that regulate the transition fromG0-G1-S-phase and G2-M. Cytokinins feature strongly as activators ofcell division in plants both in vivo and in vitro.Recent research has shown that zeatin treatment led to the up-regulationof CycD3 in Arabidopsis. This is a D-type cyclin showing stronghomology with vertebrate D cyclins which themselves are up-regulated byextracellular growth factors. Benzyladenine treatment can also shortenthe duration of S-phase through recruitment of latent origins of DNAreplication. Kinetin is involved in the phosphoregulation of the G2-Mcheckpoint; the major cyclin-dependent kinase (Cdk) at this checkpointhas recently been shown to be dephosphorylated as a result of cytokinintreatment, an effect which can also be mimicked by the fission yeastCdc25 phosphatase. Hence, a picture emerges of a cytokinin-inducedcontinuum of cell cycle activation through the up-regulation of a plantD-type cyclin at the G1 checkpoint and the phosphoregulation of the Cdkat the G2/M checkpoint. During S-phase, we argue for a link betweencytokinins and the proteins associated with replication origins.Gibberellic acid (GA) treatment induces internode elongation. Indeepwater rice, this response is mediated, at least partly, by aGA-induced up-regulation of a cyclin-Cdk at the G2-M checkpoint. Recentevidence has also linked abscisic acid to a cyclin-dependent kinaseinhibitor. These, so-called CKIs are negative regulators of Cdks whichfits with ABA's general role in growth inhibition; we await news ofethylene interactions. We highlight two instances of plant growthregulator-cell cycle interfacing during development, arguing for aninvolvement in microtubule orientation as a prerequisite to leafinitiation, and suggest a link between IAA and the activation of celldivisions in the pericycle required for lateral root initiation. A newD-type cyclin, recently discovered in Arabidopsis, may have akey role in this process. Finally, a model is presented which features ageneralised cyclin-Cdk checkpoint exhibiting various interfaces with theplant growth regulators.     

Interaction between Pseudomonas fluorescens and Meloidogyne incognita on tomato plant as influenced by the different levels of phosphorus

A pot experiment was conducted on tomato (Solanum lycopersicum cv. Pusa Ruby) to assess the effect of different phosphorus (P) levels (0, 125, 250 and 500 mg/pot) and the plant growth promoting rhizobacterium, Pseudomonas fluorescens, on the growth of tomato and on the reproduction of Meloidogyne incognita. Maximum growth of tomato occurred at P rates of 125 mg/kg soil, irrespective of whether plants were uninoculated or inoculated with P. fluorescens or M. incognita or inoculated with both the agents. Nematodes per gram of roots, egg masses per root, eggs per egg mass and galls per root significantly increased by increasing levels of P. P. fluorescens performed better than other treatments and different P levels in improving tomato growth and reducing galling and multiplication of M. incognita.     

In vitro analyses are not reliable predictors of the plant growth promotion capability of bacteria; a Pseudomonas fluorescens strain that promotes the growth and yield of wheat

Aims: In this study, we set out to identify bacteria that can be used to promote the growth of cereals, while concurrently investigating the merits of using a range of such tests to preselect bacteria for glasshouse studies. Methods and Results: A panel of 15 strains isolated from the rhizosphere and phyllosphere of cereals was tested for the ability to improve the germination of wheat seeds and for production of a range of factors associated with plant growth promotion. In parallel, all bacteria were tested for their ability to improve biomass and grain yield when applied as a soil amendment in glasshouse trials. Conclusions: There was no significant correlation between growth promotion potential in the glasshouse and the results of either the phenotypic or the germination tests. Glasshouse tests identified that only one strain, Pseudomonas fluorescens strain MKB37, gave a significant increase in head weight and grain yield. Significance and Impact of the Study: While this study has identified a candidate for further field tests, it has also highlighted the fact that the modes of action for plant growth‐promoting bacteria (PGPB) are still not fully understood, and that there is no efficient and effective screening method for identifying PGPB by laboratory tests.