Beom Ryong Kang

2R,3R-butanediol, a bacterial volatile produced by Pseudomonas chlororaphis O6, is involved in induction of systemic tolerance to drought in Arabidopsis thaliana.

Root colonization of plants with certain rhizobacteria, such as Pseudomonas chlororaphis O6, induces tolerance to biotic and abiotic stresses. Tolerance to drought was correlated with reduced water loss in P. chlororaphis O6-colonized plants and with stomatal closure, indicated by size of stomatal aperture and percentage of closed stomata. Stomatal closure and drought resistance were mediated by production of 2R,3R-butanediol, a volatile metabolite of P. chlororaphis O6. Root colonization with bacteria deficient in 2R,3R-butanediol production showed no induction of drought tolerance. Studies with Arabidopsis mutant lines indicated that induced drought tolerance required the salicylic acid (SA)-, ethylene-, and jasmonic acid-signaling pathways. Both induced drought tolerance and stomatal closure were dependent on Aba-1 and OST-1 kinase. Increases in free SA after drought stress of P. chlororaphis O6-colonized plants and after 2R,3R-butanediol treatment suggested a primary role for SA signaling in induced drought tolerance. We conclude that the bacterial volatile 2R,3R-butanediol was a major determinant in inducing resistance to drought in Arabidopsis through an SA-dependent mechanism.

Production of Indole-3-Acetic Acid in the Plant-Beneficial Strain Pseudomonas chlororaphis O6 Is Negatively Regulated by the Global Sensor Kinase GacS

Certain plant growth–promoting bacteria, such as Pseudomonas fluorescens 89B61 and Bacillus pumilus SE34, secreted high levels of indole-3-acetic acid (IAA) in tryptophan-amended medium in stationary phase as determined by chromogenic analysis and high-performance liquid chromatography. Two other growth-promoting strains, P. chlororaphis O6 and Serratia marcescens 90-166, did not produce these high levels of IAA. However, when the gacS mutant of P. chlororaphis O6 was grown in tryptophan-supplemented medium, IAA was detected in culture filtrates. IAA production by the gacS mutant in P. chlororaphis O6 was repressed in the tryptophan medium by complementation with the wild-type gacS gene. Thus, the global regulatory Gac system in P. chlororaphis O6 acts as a negative regulator of IAA production from trypophan.

Tobacco cultivars vary in induction of systemic resistance against Cucumber mosaic virus and growth promotion by Pseudomonas chlororaphis O6 and its gacS mutant

The colonization of plant roots with certain rhizosphere bacteria promotes plant growth and induces long lasting systemic protection against a broad spectrum of plant pathogens. The role of the global regulator, GacS, in the rhizosphere colonist Pseudomonas chlororaphis O6 in stimulating growth promotion and induced resistance against Cucumber mosaic virus was examined in tobacco. Responses were compared in tobacco cvs Samsun and GX3. Root colonization of Samsun with wild-type O6 and the gacS complemented mutant-elicited reduced viral symptoms and viral titre. On GX3, there was little affect on symptoms when roots were colonized by the wild-type, gacS mutant or complemented mutant but colonization by both the wild-type and the gacS mutant lowered viral titre. Wild-type O6 and the gacS mutant caused plant growth to be maintained in both tobacco cultivars after viral infection, although the affect was stronger with GX3 than Samsun. In contrast, although a chemical inducer, benzothiadiazole, reduced symptoms and viral titre in both cultivars, plant growth was suppressed. Our results indicate rhizobacteria-elicited induced viral resistance without a negative impact on growth but there was a differential response between cultivars. Detailed knowledge regarding the mechanisms inherent to these differences between cultivars requires further investigation.

Nematicidal Activity of a Nonpathogenic Biocontrol Bacterium, Pseudomonas chlororaphis O6

Bacterial culture filtrates of an aggressive rhizobacterium, Pseudomonas chlororaphis O6, displayed strong nematicidal activity. The nematicidal activity of P. chlororaphis O6 was markedly reduced in the gacS mutant of P. chlororaphis O6 grown in the presence of glycine, but no reduction of nematicidal activity in the gacS mutant was noted in the absence of glycine. The results of bioassay with P. chlororaphis O6 mutants showed that phenazine and pyrrolnitrin production was not a major factor, but the effects of glycine in the culture medium suggest that formation of hydrogen cyanide might be important. Assessments in greenhouse studies with tomatoes growing in nematode-infested soils confirmed that the application of P. chlororaphis O6 resulted in the control of the root-knot nematode. Our results demonstrated that P. chlororaphis O6 could be employed as a biocontrol agent for the control of the root-knot nematode, and the global regulator, GacS, functions as a positive regulator of the expression of nematicidal compounds and enzymes in P. chlororaphis O6.

Draft Genome Sequence of Pantoea ananatis B1-9, a Nonpathogenic Plant Growth-Promoting Bacterium

Pantoea ananatis B1-9 is an endophytic Gram-negative rhizobacterium that was isolated for its ability to promote plant growth and improve crop yield in the field. Here we report the draft genome sequence of P. ananatis B1-9. Comparison of this sequence to the sequenced genome of a plant-pathogenic P. ananatis strain, LMG20103, indicated that the pathogenesis-related genes were absent, but a subset of gene functions that may be related to its plant growth promotion were present.

Rhizobium etli USDA9032 Engineered To Produce a Phenazine Antibiotic Inhibits the Growth of Fungal Pathogens but Is Impaired in Symbiotic Performance

ABSTRACT Phenazine production was engineered in Rhizobium etli USDA9032 by the introduction of the phz locus of Pseudomonas chlororaphis O6. Phenazine-producing R. etli was able to inhibit the growth of Botrytis cinerea and Fusarium oxysporum in vitro. Black bean inoculated with phenazine-producing R. etli produced brownish Fix− nodules.

Induced Systemic Drought and Salt Tolerance by Pseudomonas chlororaphis O6 Root Colonization is Mediated by ABA-independent Stomatal Closure

Institute of Environmentally-Friendly Agriculture, Department of Plant Biotechnology, Chonnam National University, Gwangju500-757, Korea (Received on November 13, 2011; Revised on January 5, 2012; Accepted on January 11, 2012)Root colonization by the rhizobacterium Pseudomonaschlororaphis O6 in Arabidopsis thaliana Col-0 plantsresulted in induced tolerance to drought and salinitycaused by halide salt-generated ionic stress but not byosmotic stress caused by sorbitol. Stomatal aperturesdecreased following root colonization by P. chlororaphisO6 in both wild-type and ABA-insensitive Arabidopsismutant plants. These results suggest that an ABA-independent stomatal closure mechanism in the guardcells of P. chlororaphis O6-colonized plants could be akey phenotype for induced systemic tolerance todrought and salt stress. Keywords : abiotic stress, induced tolerance to environ-mental stress, priming, stomatal closurePlants possess various survival systems against abioticstresses, such as cold, drought, and salinity (Zhu, 2001).Under environmental stresses, the level of the planthormone, abscisic acid (ABA) increases triggering adaptiveresponses essential for survival (Zhu, 2001). Duringdrought and salt stresses, ABA induces stomatal closure tominimize water loss through transpiration (Leung andGiraudat, 1998). Consequently, ABA-biosynthesis mutants(aba mutants) and some of the ABA-response plantmutants (i.e. the ABA-insensitive abi mutants) aresusceptible to drought stress, due to problem of stomatalaperture regulation (Leung and Giraudat, 1998; Schroederet al., 2001). However, another plant growth regulator,jasmonate (Evans, 2003; Suhita et al., 2004), also stimu-lates stomatal closure under drought conditions (Creelmanand Mullet, 1997).Root colonization of certain plant-associated microbeselicits physiological and biochemical change to enhancesystemic resistance against various biotic and abioticstresses in plants (Kim et al., 2011; Yang et al., 2010),termed to as “induced systemic resistance (ISR)” or“induced systemic tolerance (IST)”. IST against drought orsalt stresses in plants can be induced with systemicapplication of certain rhizobacteria including Gram-positive Bacillus strains (Ryu et al., 2004; Timmusk andWagner 1999; Zhang et al., 2010), an endophytic fungalisolate, Trichoderma harziarum (Bae et al., 2010), andcertain Gram-negative bacterial isolates, such as ACC-deaminase producing bacteria (Mayak et al., 2004). Rootcolonization by Pseudomonas chlororaphis O6 inducedsystemic resistance against a broad spectrum of plantdiseases caused by viral, bacterial, and fungal pathogens invarious plants by jasmonic acid-ethylene related pathways(Cho et al., 2008; Kim et al., 2004; Ryu et al., 2007;Spencer et al., 2003). Additionally, root colonization by P.chlororaphis O6 induced systemic tol erance against drought,a process accompanied by stomatal closure. Applying2R,3R-butanediol, a volatile produced by P. chlororaphisO6, resulted in induced tolerance through a salicylic acid(SA), jasmonic acid (JA) and ethylene-dependent mech-anism (Cho et al., 2008). However, mechanisms involvedin microbe-mediated IST against drought have not beencharacterized or elucidated.In this study, we tested the hypothesis that P. chloro-raphis O6 induces tolerance to other abiotic stresses, suchas salinity, osmotic pressure, cold and heat. We used addi-tional Arabidopsis mutants altered in ABA signalingpathways to identify the role of ABA in induced abioticstress tolerance. Parental A. thaliana ecotypes Columbia(Col-0) or Landsberg erecta (Ler), and mutant and trans-genic lines were obtained from the Ohio State University

Transcriptome Analysis of Induced Systemic Drought Tolerance Elicited by Pseudomonas chlororaphis O6 in Arabidopsis thaliana.

Root colonization by Pseudomonas chlororaphis O6 induces systemic drought tolerance in Arabidopsis thaliana. Microarray analysis was performed using the 22,800-gene Affymetrix GeneChips to identify differentially-expressed genes from plants colonized with or without P. chlororaphis O6 under drought stressed conditions or normal growth conditions. Root colonization in plants grown under regular irrigation condition increased transcript accumulation from genes associated with defense, response to reactive oxygen species, and auxin- and jasmonic acid-responsive genes, but decreased transcription factors associated with ethylene and abscisic acid signaling. The cluster of genes involved in plant disease resistance were up-regulated, but the set of drought signaling response genes were down-regulated in the P. chlororaphis O6-colonized under drought stress plants compared to those of the drought stressed plants without bacterial treatment. Transcripts of the jasmonic acid-marker genes, VSP1 and pdf-1.2, the salicylic acid regulated gene, PR-1, and the ethylene-response gene, HEL, also were up-regulated in plants colonized by P. chlororaphis O6, but differed in their responsiveness to drought stress. These data show how gene expression in plants lacking adequate water can be remarkably influenced by microbial colonization leading to plant protection, and the activation of the plant defense signal pathway induced by root colonization of P. chlororaphis O6 might be a key element for induced systemic tolerance by microbes.

A Trifloxystrobin Fungicide Induces Systemic Tolerance to Abiotic Stresses

Jeonnam Agricultural Extension Service Center, Naju 520-715, Korea(Received on November 5, 2011; Revised on December 27, 2011; Accepted on January 1, 2012)Trifloxystrobin is a strobilurin fungicide, which possessesbroad spectrum control against fungal plant diseases.We demonstrated that pre-treating red pepper plantswith trifloxystrobin resulted in increased plant growthand leaf chlorophyll content compared with those incontrol plants. Relative water content of the leaves andthe survival rate of intact plants indicated that plantsacquired systemic tolerance to drought stress followingtrifloxystrobin pre-treatment. The recovery rate by re-hydration in the drought treated plant was better inthose pre-treated with trifloxystrobin than that in watertreated plants. Induced drought tolerance activity bytrifloxystrobin was sustained for 25 days after initialapplication. The trifloxystrobin treated red pepper plantsalso had induced systemic tolerance to other abioticstresses, such as frost, cold, and high temperature stresses.These findings suggest that applying the chemical fungi-cide trifloxystrobin induced systemic tolerance to cer-tain abiotic stresses in red pepper plants. Keywords : cold and freezing stresses, drought stress, hightemperature stress, induced systemic tolerance, plant growthpromotion Plants rely on many adaptive strategies against differentabiotic and biotic stresses because of their immobility.Applications of certain chemicals and plant associatedmicrobes are known to induce resistance pathways toabiotic and biotic stresses (Sticher et al., 1997; Schreiberand Desveaux, 2008; Yang et al., 2010). In systemicacquired resistance (SAR), plants develop a broad spectrumof systemic resistance against plant diseases or abioticstresses upon pathogen attack or treatment of chemicals(Ryals et al., 1996). Root colonization by biocontrolmicrobes also elicits systemic resistance, known as inducedsystemic resistance (ISR), against abiotic stresses and abroad spectrum of diseases caused by bacterial, fungal, andviral pathogens (Kim et al., 2011). Recent studies suggest that certain chemical applicationscan induce systemic resistance against abiotic or bioticstresses (Schreiber and Desveaux, 2008). Plants treatedwith one chemical inducer, β-aminobutyric acid (BABA),show dual induction for resistance to abiotic and bioticstresses (Ton et al., 2005). Application of strobilurin totobacco induces resistance against Tobacco Mosaic Virusand wildfire diseases (Herms et al., 2002). Soil applicationof a chemical pesticide, imidacloprid, reduces the incidenceof citrus canker disease (Graham and Myers, 2011). Induc-tion of drought stress tolerance in plants occurs with thefungicides, ketoconazole and propiconazole, and with theinsecticide, imidacloprid (Abdul Jaleel et al., 2007b; Han etal., 2010; Manivannan et al., 2007). Soil applications ofpropiconazole also induce tolerance to drought and inducetolerance to salinity (Adul Jallel et al., 2008), and imidaclo-prid-treated plants are tolerant to high temperature (Goniaset al., 2008). Induced tolerance to flooding and salinityoccurs with treatments with the plant regulator paclobutra-zol (Abdul Jaleel et al., 2007c; Lin et al., 2005). Trifloxystrobin, a strobilurin fungicide, is used to controla wide range of plant diseases because of its broad spectrumantifungal activity (Reuveni et al., 2000). Trifloxystrobinblocks electron transfer and inhibits mitochondrial re-spiration in the target fungus (Bartlett et al., 2002). In thisstudy, we examined whether trifloxystrobin could inducesystemic tolerance against the abiotic stresses of drought,low temperature, freezing, and high temperature in redpepper plants. We also examined the growth promotioneffects of trifloxystrobin and induced systemic resistance toplant diseases.Effect of trifloxystrobin pretreatment on plant growthand chlorophyll content. Surface-sterilized red pepperseeds were planted in pots to determine the effect of asuspension concentrate type formulation of trifloxystrobin(SC) on plant growth and photosynthesis. TrifloxystrobinSC was obtained from Bayer Cropscience Korea and was

Isolation and Characterization of Oligotrophic Bacteria Possessing Induced Systemic Disease Resistance against Plant Pathogens

Biocontrol microbes have mainly been screened among large collections of microorganisms via. nutrient-rich in vitro assays to identify novel and effective isolates. However, thus far, isolates from only a few genera, mainly spore-forming bacilli, have been commercially developed. In order to isolate field-effective biocontrol microbes, we screened for more than 200 oligotrophic bacterial strains, isolated from rhizospheres of various soil samples in Korea, which induced systemic resistance against the soft-rot disease caused by Pectobacterium carotovorum SCC1; we subsequently conducted in planta bioassay screening. Two oligotrophic bacterial strains were selected for induced systemic disease resistance against the Tobacco Mosaic Virus and the gray mold disease caused by Botrytis cinerea. The oligotrophic bacterial strains were identified as Pseudomonas manteilii B001 and Bacillus cereus C003 by biochemical analysis and the phylogenetic analysis of the 16S rRNA sequence. These bacterial strains did not exhibit any antifungal activities against plant pathogenic fungi but evidenced several other beneficial biocontrol traits, including phosphate solubilization and gelatin utilization. Collectively, our results indicate that the isolated oligotrophic bacterial strains possessing induced systemic disease resistance could provide useful tools as effective biopesticides and might be successfully used as cost-effective and preventive biocontrol agents in the field.