Physiological and genetic characterization of rice nitrogen fixer PGPR isolated from rhizosphere soils of different crops

Aims

We aimed to identify plant growth-promoting rhizobacteria that could be used to develop a biofertilizer for rice.

Methods

To obtain plant growth-promoting rhizobacteria, rhizosphere soils from different crops (rice, wheat, oats, crabgrass, maize, ryegrass, and sweet potato) were inoculated to rice plants. In total, 166 different bacteria were isolated and their plant growth-promoting traits were evaluated in terms of colony morphology, indole-3-acetic acid production, acetylene reduction activity, and phosphate solubilization activity. Moreover, genetic analysis was carried out to evaluate their phylogenetic relationships based on 16S rRNA sequence data.

Results

Strains of Bacillus altitudinis, Pseudomonas monteilii, and Pseudomonas mandelii formed associations with rice plants and fixed nitrogen. A strain of Rhizobium daejeonense showed nitrogen fixation activity in an in vitro assay and in vivo. Strains of B. altitudinis and R. daejeonense derived from rice rhizosphere soil, strains of P. monteilii and Enterobacter cloacae derived from wheat rhizosphere soil, and a strain of Bacillus pumilus derived from maize rhizosphere soil significantly promoted rice plant growth.

Conclusions

These methods are effective to identify candidate species that could be developed as biofertilizers for target crops.     

Levels of ACC and related compounds in exudate and extracts of canola seeds treated with ACC deaminase-containing plant growth-promoting bacteria

It was previously proposed that plant growth-promoting bacteria that possess 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase could utilize ACC that is present in the exudate of germinating canola seeds. The uptake and cleavage of ACC by these bacteria would lower the level of ACC, and thus ethylene within the plant, and reduce the extent of its inhibition on root elongation. To test part of the above mentioned model, ACC levels were monitored in canola seed tissues and exudate during germination. Lower amounts of ACC were present in the exudate and tissues of seeds treated with the plant growth-promoting bacterium Enterobacter cloacae CAL3, than in control seeds treated with MgSO4. The ACC-related compounds, alpha- and gamma-aminobutyric acids, both known to stimulate ethylene production, were also measured in the canola seed exudate and tissues. Approximately the same levels of alpha-aminobutyric acid were present in the exudates of the bacterium-treated seeds and the control seeds, but the amount of alpha-aminobutyric acid was lower in the tissues of the bacterium-treated seeds than in the control seeds. Smaller quantities of gamma-aminobutyric acid were seen in both the exudate and tissues of the E. cloacae CAL3-treated seeds than in the control seeds.     

An ACC Deaminase Minus Mutant of Enterobacter cloacae UW4No Longer Promotes Root Elongation

The ACC deaminase gene (acdS) from Enterobacter cloacae UW4 was replaced by homologous recombination with the acdS gene with a tetracycline resistance gene inserted within the coding region. Upon characterization of this AcdS minus mutant, it was determined that both ACC deaminase activity and the ability to promote the elongation of canola roots under gnotobiotic conditions were greatly diminished. This result is consistent with a previously postulated model that suggests that a major mechanism utilized by plant growth-promoting bacteria involves the lowering of plant ethylene levels, and hence ethylene inhibition of root elongation, by bacterial ACC deaminase. Received: 20 January 2000 / Accepted: 22 February 2000     

Involvement of gacS and rpoS in enhancement of the plant growth-promoting capabilities of Enterobacter cloacae CAL2 and UW4

The plant growth-promoting bacteria Enterobacter cloacae CAL2 and UW4 were genetically transformed with a multicopy plasmid containing an rpoS or gacS gene from Pseudomonas fluorescens. The transformed strains were compared with the nontransformed strains for growth, indoleacetic acid (IAA) production, antibiotic production, 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase activity, siderophore production, cell morphology, and the ability to promote canola root elongation. All transformed strains had a longer lag phase, were slower in reaching stationary phase, and attained a higher cell density than the nontransformed strains. Transformation resulted in cells that were significantly shorter than the nontransformed cells. The transformed strains also produced significantly more IAA than the nontransformed strains. Introduction of rpoS or gacS from Pseudomonas fluorescens was associated with a reduction in the production of both antibiotics, 2,4-diacetylphloroglucinol and mono-acetylphloroglucinol, produced by Enterobacter cloacae CAL2. With Enterobacter cloacae CAL2, plasmid-borne rpoS, but not gacS, increased the level of ACC deaminase activity, while introduction of rpoS in Enterobacter cloacae UW4 caused a decrease in ACC deaminase activity. Neither gacS nor rpoS significantly affected the level of siderophores synthesized by either bacterial strain. Overproduction of either GacA or RpoS in Enterobacter cloacae CAL2 resulted in a significant increase in the root lengths of canola seedlings when seeds were treated with the bacteria, and overproduction of RpoS caused an increase in canola shoot as well as root lengths.     

Preliminary investigations on inducing salt tolerance in maize through inoculation with rhizobacteria containing ACC deaminase activity

Twenty rhizobacterial strains containing 1-aminocyclopropane-1-carboxylate deaminase were isolated from the rhizosphere of salt-affected maize fields. They were screened for their growth-promoting activities under axenic conditions at 1, 4, 8, and 12 dS x m-1 salinity levels. Based upon the data of the axenic study, the 6 most effective strains were selected to conduct pot trials in the wire house. Besides one original salinity level (1.6 dS x m-1), 3 other salinity levels (4, 8, and 12 dS x m-1) were maintained in pots and maize seeds inoculated with selected strains of plant growth-promoting rhizobacteria, as well as uninoculated controls were sown. Results showed that the increase in salinity level decreased the growth of maize seedlings. However, inoculation with rhizobacterial strains reduced this depression effect and improved the growth and yield at all the salinity levels tested. Selected strains significantly increased plant height, root length, total biomass, cob mass, and grain yield up to 82%, 93%, 51%, 40%, and 50%, respectively, over respective uninoculated controls at the electrical conductivity of 12 dS x m-1. Among various plant growth-promoting rhizobacterial strains, S5 (Pseudomonas syringae), S14 (Enterobacter aerogenes), and S20 (Pseudomonas fluorescens) were the most effective strains for promoting the growth and yield of maize, even at high salt stress. The relatively better salt tolerance of inoculated plants was associated with a high K+/Na+ ratio as well as high relative water and chlorophyll and low proline contents.     

Canopy photosynthesis modulates soil respiration in a temperate semi-arid shrubland at multiple timescales

Plant and Soil - Plant growth-promoting bacteria of the genus Bacillus are known to solubilize phosphates and enhance plant growth in many plant species. We explored the effects of the inoculation...     

Plant growth enhancement and associated physiological responses are coregulated by ethylene and gibberellin in response to harpin protein Hpa1

The harpin protein Hpa1 produced by the bacterial blight pathogen of rice induces several growth-promoting responses in plants, activating the ethylene signaling pathway, increasing photosynthesis rates and EXPANSIN (EXP) gene expression levels, and thereby enhancing the vegetative growth. This study was attempted to analyze any mechanistic connections among the above and the role of gibberellin in these responses. Hpa1-induced growth enhancement was evaluated in Arabidopsis, tomato, and rice. And growth-promoting responses were determined mainly as an increase of chlorophyll a/b ratio, which indicates a potential elevation of photosynthesis rates, and enhancements of photosynthesis and EXP expression in the three plant species. In Arabidopsis, Hpa1-induced growth-promoting responses were partially compromised by a defect in ethylene perception or gibberellin biosynthesis. In tomato and rice, compromises of Hpa1-induced growth-promoting responses were caused by a pharmacological treatment with an ethylene perception inhibitor or a gibberellin biosynthesis inhibitor. In the three plant species, moreover, Hpa1-induced growth-promoting responses were significantly impaired, but not totally eliminated, by abolishing ethylene perception or gibberellin synthesis. However, simultaneous nullifications in both ethylene perception and gibberellin biosynthesis almost canceled the full effects of Hpa1 on plant growth, photosynthesis, and EXP2 expression. Theses results suggest that ethylene and gibberellin coregulate Hpa1-induced plant growth enhancement and associated physiological and molecular responses.     

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.     

田菁根际促生菌的筛选及其促生耐盐效果

【目的】为探究盐生植物田菁及其根际功能微生物改良盐碱地的效果,本研究从黄河三角洲盐碱区田菁根际土壤中分离促生菌,并明确其耐盐促生效果。【方法】采用选择培养方法从田菁根际土壤中分离固氮菌、解磷菌以及解钾菌,并进行16S rRNA分子生物学鉴定。之后对菌株的耐盐及促生特性进行测定,筛选性状优良菌株进行玉米促生作用研究。【结果】共分离得到105株根际促生菌,其中N102兼具多种促生特性且耐盐性达15%。田菁种子发芽试验表明,N102可显著提高田菁发芽率(47%,P<0.05)、芽长(48.5%,P<0.05)和根长(60%,P<0.05);玉米盆栽试验结果表明,N102对盐胁迫下玉米的株高、根长、叶绿素含量、地上部干重以及根干重具有显著的促进作用。经系统发育分析,N102与Enterobacter soli ATCC BAA-2102 (NR117547)序列相似度为99.30%,鉴定属于Enterobacter属。【结论】菌株N102具有多种植物促生耐盐特性,具有开发成有效促进盐碱地作物生长的微生物肥的良好前景。     

Draft genome sequence of Paenibacillus peoriae strain KCTC 3763T

Paenibacillus peoriae is a potentially plant-beneficial soil bacterium and is a close relative to Paenibacillus polymyxa, the type species of the genus Paenibacillus. Herein, we present the 5.77-Mb draft genome sequence of the P. peoriae type strain with the aim of providing insight into the genomic basis of plant growth-promoting Paenibacillus species.     

Co-Inoculation of Common Bean with Rhizobium and Azospirillum Enhance the Drought Tolerance

Russian Journal of Plant Physiology - Plant growth-promoting rhizobacteria (PGPR) have been shown to reduce the negative effects of drought on numerous plant species, but these effects are unknown...     

Joint Immobilization of Plant Growth-Promoting Bacteria and Green Microalgae in Alginate Beads as an Experimental Model for Studying Plant-Bacterium Interactions

A simple, quantitative experimental model, offering a convenient and basic approach to studies of plant-bacterium interactions, is proposed. This involves immobilizing a unicellular, freshwater microalga, a Chlorella species, serving as the plant, with a plant growth-promoting bacterium, an Azospirillum species, in small alginate beads to allow close interaction and to avoid external interference from bacterial contaminants.     

乙偶姻生物合成代谢调控及其应用

许多微生物在糖酵解过程中能够将糖类转化为乙偶姻来避免过度酸化。乙偶姻能够调节NAD＋/NADH的比率并存储碳源。此外,乙偶姻作为一种具有特殊奶油香气的食用香料,广泛应用于食品,烟草、酒类和化妆品行业。近些年研究发现许多植物根际促生菌能够通过产生乙偶姻激活植物对外界环境压力的抗性,激活植物系统抗性,抵御病原菌的侵袭。乙偶姻还可以促进植物生长,提高产量。另外,乙偶姻还是调节根际促生菌与宿主植物相互作用的信号分子。简述乙偶姻的生物合成路径及其调控,并介绍乙偶姻在食品、医药、化工、化妆品、植物保护、生物燃料等方面的应用。     

Characterization of plant growth-promoting bacteria associated with rice cropped in iron-stressed soils

Plant growth-promoting rhizobacteria (PGPR) are able to promote plant growth using a wide variety of mechanisms as well as provide bioprotection against biotic and abiotic stresses. The objectives of this study were to isolate and characterize putative PGPR associated with rice cultivars with a distinct tolerance to iron toxicity grown in two areas: one area with a well-established history of iron toxicity and another without iron toxicity. Bacterial strains were selectively isolated based on their growth in selective media and were identified by partial sequencing of their 16S rRNA genes. Bacterial isolates were evaluated for their ability to produce indolic compounds, siderophores, and ACC deaminase and to solubilize tricalcium phosphates. In vitro biological nitrogen fixation was evaluated for the bacterial isolates used in the inoculation experiments. A total of 329 bacterial strains were isolated. The composition of the bacterial genera and the occurrence of different plant growth-promoting (PGP) traits were significantly affected by the iron conditions and by the cultivar. Strains belonging to the Burkholderia and Enterobacter genera were the most abundant of all the Gram-negative isolates, and those belonging to the Paenibacillus and Bacillus genera were the most abundant of the Gram-positive isolates. A large number of putative PGPR belonging to different bacterial genera presented several PGP traits. Strains belonging to the Burkholderia, Chryseobacterium, and Ochrobactrum genera contributed to plant growth as well as to enhanced nutrient uptake of the rice plants in in vivo experiments. Growth and nutrient uptake of plants inoculated with isolate FeS53 (Paenibacillus sp.) in the presence of an iron excess were similar to those of plants submitted to the control iron condition, indicating that this bacterium can mitigate the effects caused by iron stress.     

Bacterial contaminants of micropropagated plant cultures

Bacterial contaminants of micropropagated plant cultures were isolated and characterized with standard bacteriological tests and appropriate API strips. Results obtained were analysed by the API identification software. Of 198 bacterial strains isolated from nine plant species, 90% were identified as Bacillus, Enterobacter, Micrococcus, Staphylococcus, Pseudomonas or Lactobacillus species. Possible sources of contamination are discussed.     

<Emphasis Type="Italic">Herbaspirillum</Emphasis> Species: A Potential Pathogenic Bacteria Isolated from Acute Lymphoblastic Leukemia Patient

Herbaspirillum species, colonized the plant rhizosphere, also called rhizobacteria, are plant growth-promoting bacteria. Recently we isolated Herbaspirillum from blood cultures of acute lymphoblastic leukemia (ALL) and identified by PCR and gene sequencing. Herbaspirillum may be a potential pathogenic bacteria. Although the exact role that these species play in ALL patients is unknown, their differentiation from other species has serious implications for clinical care and patient well-being.     

Promotion of Plant Growth by Bacterial ACC Deaminase

To date, there has been only limited commercial use of plant growth-promoting bacteria in agriculture, horticulture, and silviculture. However, with recent progress toward understanding the mechanisms that these organisms utilize to facilitate plant growth, the use of plant growth-promoting bacteria is expected to continue to increase worldwide. One of the key mechanisms employed by plant growth-promoting bacteria to facilitate plant growth is the lowering of plant ethylene levels by the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase. This article reviews the published work on this enzyme, with an emphasis on its biochemistry, protein structure, genes, and regulation. In addition, this article provides some initial insights into the changes in both plants and ACC deaminase-containing plant growth-promoting bacteria as a consequence of plant-microbe interactions. Finally, a brief discussion of how bacterial ACC deaminase and indoleacetic acid (IAA) together modulate plant growth and development is included.     

Tomato growth and resistance promotion by Enterobacter hormaechei subsp. steigerwaltii EB8D

Fusarium wilt is an economically important disease of tomato crop (Solanum lycopersicum L). Plant growth-promoting rhizobacteria (PGPR) represent an alternative to improve plant growth and yield as well as to act as agent of biocontrol. In this study, antagonistic effects of four selected isolates (EB8D, EB20J, EB24L and EB26M) were evaluated against Fusarium oxysporum f.sp. radicis lycopersici (FORL) as potential biocontrol agents in vitro and in vivo. After 30?days of culture, dry weight, length of stem and root were significantly (p?≤?.05) higher compared with the non-inoculated control. Compared with the control plate (inoculated only with pathogen), EB8D had showed efficient antagonism against FORL (48.88%). The different strains have been screened for siderophore production, solubilisation of mineral phosphates, synthesis of indolic acetic acid (IAA) to show the plant growth-promoting potential. The experimental groups were compared with a control group that did not receive any treatment by FORL, and EB8D was the best isolate in terms of growth promotion with an improvement of 73.85% of the stem length, 110.86% of the root length and 118.85% of the dry weight comparing with the non-treated controls. Compared with a control group treated by FORL, biocontrol activity has shown that EB8D strain improved the stem length with 111.85%, the root length with 118.85% and the dry weight with 452.38%. 16S rRNA analysis has confirmed that this strain belongs to the genus Enterobacter and has high similarity with Enterobacter hormaechei subsp. steigerwaltii (99.71%). EB8D has a significant strong protective potential against FORL and lead to better tomato growth and might have biotechnological potential for controlling fusarium wilt in tomato plants.     

Enterobacter radicincitans sp. nov., a plant growth promoting species of the family Enterobacteriaceae

A plant growth promoting bacterial isolate (D5/23T) from the phyllosphere of winter wheat, able to fix atmospheric nitrogen and to produce auxines and cytokinins was investigated in a polyphasic taxonomy approach. Phylogenetic analyses using the 16S rRNA gene sequence of the strain clearly indicated that the strain belonged to the family Enterobacteriaceae, most closely related to Enterobacter cloacae with 99.0% and Enterobacter dissolvens with 98.5% sequence similarity. Phylogenetic analysis derived from the sequence of the rpoB gene showed the highest sequence similarities to Enterobacter cowanii (93.0%) but supported the distinct position of strain D5/23T. The isolate produced a fatty acid pattern typical for members of the family Enterobacteriaceae. On the basis of the phylogenetic analyses, DNA-DNA hybridizations, and the unique physiological and biochemical characteristics, we propose that strain D5/23T represents a new species of the genus Enterobacter for which we propose the name Enterobacter radicincitans sp. nov.     

Revegetation of a lakeside barren area by the application of plant growth-promoting rhizobacteria

The growth stimulation of wild plants by several bacterial species showing plant growth-promoting capabilities was examined in a barren lakeside area at Lake Paro, Korea. Microbial numbers and activities in the field soil were monitored for 73 days after inoculation of the bacteria. The acridine orange direct counts for the total soil bacterial populations ranged between 2.0-2.3x10(9) cells/g soil and 1.4-1.8x10(9) cells/g soil in the inoculated and uninoculated soils, respectively. The numbers of Pseudomonas spp., which is known as a typical plant growth-promoting rhizobacteria, and the total microbial activity were higher in the inoculated soil compared to those in the uninoculated soil. The average shoot and root lengths of the wild plants grown in the inoculated soil were 17.3 cm and 12.4 cm, respectively, and longer than those of 11.4 cm and 8.5 cm in the uninoculated soil. The total dry weight of the harvested wild plants was also higher in the inoculated soil (42.0 g) compared to the uninoculated soil (35.1 g). The plant growth-promoting capabilities of the inoculated bacteria may be used for the rapid revegetation of barren or disturbed land, and as biofertilizer in agriculture.