[Show abstract][Hide abstract] ABSTRACT: The aim of this study was to inventory the types of plant growth-promoting rhizobacteria (PGPR) present in the rhizosphere of plants grown in soils contaminated with heavy metals, recalcitrant organics, petroleum sewage or salinity in China. We screened 1223 isolates for antifungal activity and about 24% inhibited Rhizoctonia solani or Sclerotinia sclerotiorum. Twenty-four strains inhibitory to R. solani, Gaeumannomyces graminis var. tritici and/or S. sclerotiorum and representing the dominant morphotypes were assayed for PGPR activity. Seven strains contained phlD, prnD, pltC or phzF genes and produced the antibiotics 2,4-diacetylphloroglucinol, pyrrolnitrin, pyoluteorin and phenazines respectively. Six strains contained acdS, which encodes 1-aminocyclopropane-1-carboxylic acid deaminase. Phylogenetic analysis of 16S rDNA and phlD, phzF and acdS genes demonstrated that some strains identified as Pseudomonas were similar to model PGPR strains Pseudomonas protegens Pf-5, Pseudomonas chlororaphis subsp. aureofaciens 30–84 and P. brassicacearum Q8r1-96. Pseudomonas protegens- and P. chlororaphis-like strains had the greatest biocontrol activity against Rhizoctonia root rot and take-all of wheat. Pseudomonas protegens and P. brassicacearum-like strains showed the greatest promotion of canola growth. Our results indicate that strains from contaminated soils are similar to well-described PGPR found in agricultural soils worldwide.
[Show abstract][Hide abstract] ABSTRACT: Beneficial microbes in the microbiome of plant roots improve plant health. Induced systemic resistance (ISR) emerged as an important mechanism by which selected plant growth-promoting bacteria and fungi in the rhizosphere prime the whole plant body for enhanced defense against a broad range of pathogens and insect herbivores. A wide variety of root-associated mutualists, including Pseudomonas, Bacillus, Trichoderma, and mycorrhiza species sensitize the plant immune system for enhanced defense without directly activating costly defenses. This review focuses on molecular processes at the interface between plant roots and ISR-eliciting mutualists, and on the progress in our understanding of ISR signaling and systemic defense priming. The central role of the root-specific transcription factor MYB72 in the onset of ISR and the role of phytohormones and defense regulatory proteins in the expression of ISR in aboveground plant parts are highlighted. Finally, the ecological function of ISR-inducing microbes in the root microbiome is discussed. Expected final online publication date for the Annual Review of Phytopathology Volume 52 is August 04, 2014. Please see http://www.annualreviews.org/catalog/pubdates.aspx for revised estimates.
Annual Review of Phytopathology 09/2014; 52:347-375. · 10.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: ABSTRACT Pseudomonas fluorescens HC1-07, previously isolated from the phyllosphere of wheat grown in Hebei province, China, suppresses the soilborne disease of wheat take-all, caused by Gaeumannomyces graminis var. tritici. We report here that strain HC1-07 also suppresses Rhizoctonia root rot of wheat caused by Rhizoctonia solani AG-8. Strain HC1-07 produced a cyclic lipopeptide (CLP) with a molecular weight of 1,126.42 based on analysis by electrospray ionization mass spectrometry. Extracted CLP inhibited the growth of G. graminis var. tritici and R. solani in vitro. To determine the role of this CLP in biological control, plasposon mutagenesis was used to generate two nonproducing mutants, HC1-07viscB and HC1-07prtR2. Analysis of regions flanking plasposon insertions in HC1-07prtR2 and HC1-07viscB revealed that the inactivated genes were similar to prtR and viscB, respectively, of the well-described biocontrol strain P. fluorescens SBW25 that produces the CLP viscosin. Both genes in HC1-07 were required for the production of the viscosin-like CLP. The two mutants were less inhibitory to G. graminis var. tritici and R. solani in vitro and reduced in ability to suppress take-all. HC1-07viscB but not HC-07prtR2 was reduced in ability to suppress Rhizoctonia root rot. In addition to CLP production, prtR also played a role in protease production.
[Show abstract][Hide abstract] ABSTRACT: In agro-ecosystems worldwide, some of the most important and devastating diseases are caused by soil-borne necrotrophic fungal pathogens, against which crop plants generally lack genetic resistance. However, plants have evolved approaches to protect themselves against pathogens by stimulating and supporting specific groups of beneficial microorganisms that have the ability to protect either by direct inhibition of the pathogen or by inducing resistance mechanisms in the plant. One of the best examples of protection of plant roots by antagonistic microbes occurs in soils that are suppressive to take-all disease of wheat. Take-all, caused by Gaeumannomyces graminis var. tritici, is the most economically important root disease of wheat worldwide. Take-all decline (TAD) is the spontaneous decline in incidence and severity of disease after a severe outbreak of take-all during continuous wheat or barley monoculture. TAD occurs worldwide, and in the United States and The Netherlands it results from a build-up of populations of 2,4-diacetylphloroglucinol (2,4-DAPG)-producing fluorescent Pseudomonas spp. during wheat monoculture. The antibiotic 2,4-DAPG has a broad spectrum of activity and is especially active against the take-all pathogen. Based on genotype analysis by repetitive sequence-based-PCR analysis and restriction fragment length polymorphism of phlD, a key 2,4-DAPG biosynthesis gene, at least 22 genotypes of 2,4-DAPG producing fluorescent Pseudomonas spp. have been described worldwide. In this review, we provide an overview of G. graminis var. tritici, the take-all disease, Pseudomonas biocontrol agents, and mechanism of disease suppression.
The plant pathology journal. 06/2013; 29(2):125-35.
[Show abstract][Hide abstract] ABSTRACT: We investigated the taxonomic placement of phenazine-producing fluorescent Pseudomonas spp. in the Inland Pacific Northwest, USA. Five distinct species were identified, two of which were provisionally considered to be new. Agroclimatic zone and soil silt content affected the species diversity across the region.
Applied and Environmental Microbiology 04/2013; · 3.95 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: BACKGROUND:Yeast deletion-mutant collections have been successfully used to infer the mode-of-action of drugs especially by profiling chemical-genetic and genetic-genetic interactions on a genome-wide scale. Although tens of thousands of those profiles are publicly available, a lack of an accurate method for mining such data has been a major bottleneck for more widespread use of these useful resources.RESULTS:For general usage of those public resources, we designed FitRankDB as a general repository of fitness profiles, and developed a new search algorithm, FitSearch, for identifying the profiles that have a high similarity score with statistical significance for a given fitness profile. We demonstrated that our new repository and algorithm are highly beneficial to researchers who attempting to make hypotheses based on unknown modes-of-action of bioactive compounds, regardless of the types of experiments that have been performed using yeast deletion-mutant collection in various types of different measurement platforms, especially non-chip-based platforms.CONCLUSIONS:We showed that our new database and algorithm are useful when attempting to construct a hypothesis regarding the unknown function of a bioactive compound through small-scale experiments with a yeast deletion collection in a platform independent manner. The FitRankDB and FitSearch enhance the ease of searching public yeast fitness profiles and obtaining insights into unknown mechanisms of action of drugs. FitSearch is freely available at http://fitsearch.kaist.ac.kr.
[Show abstract][Hide abstract] ABSTRACT: Phenazine compounds represent a large class of bacterial metabolites that are produced by some fluorescent Pseudomonas spp. and a few other bacterial genera. Phenazines were first noted in the scientific literature over 100 years ago, but for a long time were considered to be pigments of uncertain function. Following evidence that phenazines act as virulence factors in the opportunistic human and animal pathogen Pseudomonas aeruginosa and are actively involved in the suppression of plant pathogens, interest in these compounds has broadened to include investigations of their genetics, biosynthesis, activity as electron shuttles, and contribution to the ecology and physiology of the cells that produce them. This minireview highlights some recent and exciting insights into the diversity, frequency and ecological roles of phenazines produced by fluorescent Pseudomonas spp.
[Show abstract][Hide abstract] ABSTRACT: We provide here a comparative genome analysis of ten strains within the Pseudomonas fluorescens group including seven new genomic sequences. These strains exhibit a diverse spectrum of traits involved in biological control and other multitrophic interactions with plants, microbes, and insects. Multilocus sequence analysis placed the strains in three sub-clades, which was reinforced by high levels of synteny, size of core genomes, and relatedness of orthologous genes between strains within a sub-clade. The heterogeneity of the P. fluorescens group was reflected in the large size of its pan-genome, which makes up approximately 54% of the pan-genome of the genus as a whole, and a core genome representing only 45-52% of the genome of any individual strain. We discovered genes for traits that were not known previously in the strains, including genes for the biosynthesis of the siderophores achromobactin and pseudomonine and the antibiotic 2-hexyl-5-propyl-alkylresorcinol; novel bacteriocins; type II, III, and VI secretion systems; and insect toxins. Certain gene clusters, such as those for two type III secretion systems, are present only in specific sub-clades, suggesting vertical inheritance. Almost all of the genes associated with multitrophic interactions map to genomic regions present in only a subset of the strains or unique to a specific strain. To explore the evolutionary origin of these genes, we mapped their distributions relative to the locations of mobile genetic elements and repetitive extragenic palindromic (REP) elements in each genome. The mobile genetic elements and many strain-specific genes fall into regions devoid of REP elements (i.e., REP deserts) and regions displaying atypical tri-nucleotide composition, possibly indicating relatively recent acquisition of these loci. Collectively, the results of this study highlight the enormous heterogeneity of the P. fluorescens group and the importance of the variable genome in tailoring individual strains to their specific lifestyles and functional repertoire.
[Show abstract][Hide abstract] ABSTRACT: Pseudomonas fluorescens 2112, isolated in Korea as an indigenous antagonistic bacteria, can produce 2,4- diacetylphloroglucinol (2,4-DAPG) and the siderophore pyoveridin2112 for the control of phytophthora blight of red-pepper. P. fluorescens 2112 was classified into a new genotype C among the 17 genotypes of 2,4-DAPG producers, by phlD restriction fragment length polymorphism (RFLP). The colonizing ability of P. fluorescens 2112 in pea rhizosphere was equal to the well-known pea colonizers, P. fluorescens Q8r1 (genotype D) and MVP1-4 (genotype P), after 6 cycling cultivations for 18 weeks. Four tested 2,4- DAPG-producing Pseudomonas spp. could colonize with about a 96% dominance ratio against total bacteria in pea rhizosphere. The strain P. fluorescens 2112 was as good a colonizer as other Pseudomonas spp. genotypes in pea plant growth-promoting rhizobacteria.
Journal of Microbiology and Biotechnology 06/2012; 22(6):763-70. · 1.40 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Pseudomonas fluorescens strains that produce the polyketide antibiotic 2,4-diacetylphloroglucinol (2,4-DAPG) are among the most effective rhizobacteria that suppress root and crown rots, wilts, and damping-off diseases of a variety of crops, and they play a key role in the natural suppressiveness of some soils to certain soilborne pathogens. Root colonization by 2,4-DAPG-producing P. fluorescens strains Pf-5 (genotype A), Q2-87 (genotype B), Q8r1-96 (genotype D), and HT5-1 (genotype N) produced induced systemic resistance (ISR) in Arabidopsis thaliana accession Col-0 against bacterial speck caused by P. syringae pv. tomato. The ISR-eliciting activity of the four bacterial genotypes was similar, and all genotypes were equivalent in activity to the well-characterized strain P. fluorescens WCS417r. The 2,4-DAPG biosynthetic locus consists of the genes phlHGF and phlACBDE. phlD or phlBC mutants of Q2-87 (2,4-DAPG minus) were significantly reduced in ISR activity, and genetic complementation of the mutants restored ISR activity back to wild-type levels. A phlF regulatory mutant (overproducer of 2,4-DAPG) had ISR activity equivalent to the wild-type Q2-87. Introduction of DAPG into soil at concentrations of 10 to 250 μM 4 days before challenge inoculation induced resistance equivalent to or better than the bacteria. Strain Q2-87 induced resistance on transgenic NahG plants but not on npr1-1, jar1, and etr1 Arabidopsis mutants. These results indicate that the antibiotic 2,4-DAPG is a major determinant of ISR in 2,4-DAPG-producing P. fluorescens, that the genotype of the strain does not affect its ISR activity, and that the activity induced by these bacteria operates through the ethylene- and jasmonic acid-dependent signal transduction pathway.
[Show abstract][Hide abstract] ABSTRACT: Certain strains of the rhizosphere bacterium Pseudomonas fluorescens contain the phenazine biosynthesis operon (phzABCDEFG) and produce redox-active phenazine antibiotics that suppress a wide variety of soilborne plant pathogens. In 2007 and 2008, we isolated 412 phenazine-producing (Phz(+)) fluorescent Pseudomonas strains from roots of dryland wheat and barley grown in the low-precipitation region (<350 mm annual precipitation) of central Washington State. Based on results of BOX-PCR genomic fingerprinting analysis, these isolates, as well as the model biocontrol Phz(+) strain P. fluorescens 2-79, were assigned to 31 distinct genotypes separated into four clusters. All of the isolates exhibited high 16S rDNA sequence similarity to members of the P. fluorescens species complex including Pseudomonas orientalis, Pseudomonas gessardii, Pseudomonas libanensis, and Pseudomonas synxantha. Further recA-based sequence analyses revealed that the majority of new Phz(+) isolates (386 of 413) form a clade distinctly separated from P. fluorescens 2-79. Analysis of phzF alleles, however, revealed that the majority of those isolates (280 of 386) carried phenazine biosynthesis genes similar to those of P. fluorescens 2-79. phzF-based analyses also revealed that phenazine genes were under purifying selection and showed evidence of intracluster recombination. Phenotypic analyses using Biolog substrate utilization and observations of phenazine-1-carboxylic acid production showed considerable variability amongst members of all four clusters. Biodiversity indices indicated significant differences in diversity and evenness between the sampled sites. In summary, this study revealed a genotypically and phenotypically diverse group of phenazine producers with a population structure not seen before in indigenous rhizosphere-inhabiting Phz(+) Pseudomonas spp.
[Show abstract][Hide abstract] ABSTRACT: This work determined the impact of irrigation on the seasonal dynamics of populations of Pseudomonas spp. producing the antibiotics phenazine-1-carboxylic acid (Phz(+)) and 2,4-diacetylphloroglucinol (Phl(+)) in the rhizosphere of wheat grown in the low-precipitation zone (150 to 300 mm annually) of the Columbia Plateau of the Inland Pacific Northwest. Population sizes and plant colonization frequencies of Phz(+) and Phl(+) Pseudomonas spp. were determined in winter and spring wheat collected during the growing seasons from 2008 to 2009 from selected commercial dryland and irrigated fields in central Washington State. Only Phz(+) bacteria were detected on dryland winter wheat, with populations ranging from 4.8 to 6.3 log CFU g(-1) of root and rhizosphere colonization frequencies of 67 to 100%. The ranges of population densities of Phl(+) and Phz(+) Pseudomonas spp. recovered from wheat grown under irrigation were similar, but 58 to 100% of root systems were colonized by Phl(+) bacteria whereas only 8 to 50% of plants harbored Phz(+) bacteria. In addition, Phz(+) Pseudomonas spp. were abundant in the rhizosphere of native plant species growing in nonirrigated areas adjacent to the sampled dryland wheat fields. This is the first report that documents the impact of irrigation on indigenous populations of two closely related groups of antibiotic-producing pseudomonads that coinhabit the rhizosphere of an economically important cereal crop. These results demonstrate how crop management practices can influence indigenous populations of antibiotic-producing pseudomonads with the capacity to suppress soilborne diseases of wheat.
Applied and Environmental Microbiology 03/2012; 78(9):3214-20. · 3.95 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Fluorescent Pseudomonas isolated from the rhizosphere of diverse plants have been studied as biocontrol agents of soilborne pathogens worldwide.
Certain strains of these bacteria are capable of exerting a variety of mechanisms of plant growth promotion and protection,
including the production of the natural antibiotics 2,4-diacetylphloroglucinol (2,4-DAPG) and derivatives of phenazine (Phz).
These compounds are broadly active against plant pathogens and are produced by widely distributed taxonomically diverse Pseudomonas spp. that inhabit the rhizosphere of cereal crops and render certain soils naturally suppressive to soilborne plant diseases.
There is mounting evidence that the role of 2,4-DAPG and phenazines in the rhizosphere is not limited to antibiosis but also
involves regulatory and signaling functions, induction of systemic resistance, and reduction of minerals in soil.
[Show abstract][Hide abstract] ABSTRACT: Natural antibiotics are thought to function in the defense, fitness, competitiveness, biocontrol activity, communication, and gene regulation of microorganisms. However, the scale and quantitative aspects of antibiotic production in natural settings are poorly understood. We addressed these fundamental questions by assessing the geographic distribution of indigenous phenazine-producing (Phz(+)) Pseudomonas spp. and the accumulation of the broad-spectrum antibiotic phenazine-1-carboxylic acid (PCA) in the rhizosphere of wheat grown in the low-precipitation zone (<350 mm) of the Columbia Plateau and in adjacent, higher-precipitation areas. Plants were collected from 61 commercial wheat fields located within an area of about 22,000 km(2). Phz(+) Pseudomonas spp. were detected in all sampled fields, with mean population sizes ranging from log 3.2 to log 7.1 g(-1) (fresh weight) of roots. Linear regression analysis demonstrated a significant inverse relationship between annual precipitation and the proportion of plants colonized by Phz(+) Pseudomonas spp. (r(2) = 0.36, P = 0.0001). PCA was detected at up to nanomolar concentrations in the rhizosphere of plants from 26 of 29 fields that were selected for antibiotic quantitation. There was a direct relationship between the amount of PCA extracted from the rhizosphere and the population density of Phz(+) pseudomonads (r(2) = 0.46, P = 0.0006). This is the first demonstration of accumulation of significant quantities of a natural antibiotic across a terrestrial ecosystem. Our results strongly suggest that natural antibiotics can transiently accumulate in the plant rhizosphere in amounts sufficient not only for inter- and intraspecies signaling but also for the direct inhibition of sensitive organisms.
Applied and Environmental Microbiology 12/2011; 78(3):804-12. · 3.95 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Take-all disease of wheat caused by the soilborne fungus Gaeumannomyces graminis var. tritici is one of the most important root diseases of wheat worldwide. Bacteria were isolated from winter wheat from irrigated and rainfed fields in Hebei and Jiangsu provinces in China, respectively. Samples from rhizosphere soil, roots, stems, and leaves were plated onto King's medium B agar and 553 isolates were selected. On the basis of in vitro tests, 105 isolates (19% of the total) inhibited G. graminis var. tritici and all were identified as Pseudomonas spp. by amplified ribosomal DNA restriction analysis. Based on biocontrol assays, 13 strains were selected for further analysis. All of them aggressively colonized the rhizosphere of wheat and suppressed take-all. Of the 13 strains, 3 (HC9-07, HC13-07, and JC14-07, all stem endophytes) had genes for the biosynthesis of phenazine-1-carboxylic acid (PCA) but none had genes for the production of 2,4-diacetylphloroglucinol, pyoluteorin, or pyrrolnitrin. High-pressure liquid chromatography (HPLC) analysis of 2-day-old cultures confirmed that HC9-07, HC13-07, and JC14-07 produced PCA but no other phenazines were detected. HPLC quantitative time-of-flight 2 mass-spectrometry analysis of extracts from roots of spring wheat colonized by HC9-07, HC13-07, or Pseudomonas fluorescens 2-79 demonstrated that all three strains produced PCA in the rhizosphere. Loss of PCA production by strain HC9-07 resulted in a loss of biocontrol activity. Analysis of DNA sequences within the key phenazine biosynthesis gene phzF and of 16S rDNA indicated that strains HC9-07, HC13-07, and JC14-07 were similar to the well-described PCA producer P. fluorescens 2-79. This is the first report of 2-79-like bacteria being isolated from Asia.
[Show abstract][Hide abstract] ABSTRACT: 2,4-Diacetylphloroglucinol (2,4-DAPG), an antibiotic produced by Pseudomonas fluorescens, has broad-spectrum antibiotic activity, inhibiting organisms ranging from viruses, bacteria, and fungi to higher plants and mammalian cells. The biosynthesis and regulation of 2,4-DAPG in P. fluorescens are well described, but the mode of action against target organisms is poorly understood. As a first step to elucidate the mechanism, we screened a deletion library of Saccharomyces cerevisiae in broth and agar medium supplemented with 2,4-DAPG. We identified 231 mutants that showed increased sensitivity to 2,4-DAPG under both conditions, including 22 multidrug resistance-related mutants. Three major physiological functions correlated with an increase in sensitivity to 2,4-DAPG: membrane function, reactive oxygen regulation, and cell homeostasis. Physiological studies with wild-type yeast validated the results of the mutant screens. The chemical-genetic fitness profile of 2,4-DAPG resembled those of menthol, sodium azide, and hydrogen peroxide determined in previous high-throughput screening studies. Collectively, these findings indicate that 2,4-DAPG acts on multiple basic cellular processes.
Applied and Environmental Microbiology 12/2010; 77(5):1770-6. · 3.95 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Pseudomonas fluorescens Q8r1-96 represents a group of rhizosphere strains responsible for the suppressiveness of agricultural soils to take-all disease of wheat. It produces the antibiotic 2,4-diacetylphloroglucinol and aggressively colonizes the roots of cereal crops. In this study, we analyzed the genome of Q8r1-96 and identified a type III protein secretion system (T3SS) gene cluster that has overall organization similar to that of the T3SS gene cluster of the plant pathogen Pseudomonas syringae. We also screened a collection of 30 closely related P. fluorescens strains and detected the T3SS genes in all but one of them. The Q8r1-96 genome contained ropAA and ropM type III effector genes, which are orthologs of the P. syringae effector genes hopAA1-1 and hopM1, as well as a novel type III effector gene designated ropB. These type III effector genes encoded proteins that were secreted in culture and injected into plant cells by both P. syringae and Q8r1-96 T3SSs. The Q8r1-96 T3SS was expressed in the rhizosphere, but mutants lacking a functional T3SS were not altered in their rhizosphere competence. The Q8r1-96 type III effectors RopAA, RopB, and RopM were capable of suppressing the hypersensitive response and production of reactive oxygen species, two plant immune responses.
Journal of bacteriology 10/2010; 193(1):177-89. · 3.94 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Dark pigmented fungi of the Gaeumannomyces-Phialophora complex were isolated from the roots of wheat grown in fields in eastern Washington State. These fungi were identified as Phialophora spp. on the basis of morphological and genetic characteristics. The isolates produced lobed hyphopodia on wheat coleoptiles, phialides, and hyaline phialospores. Sequence comparison of internal transcribed spacer regions indicated that the Phialophora isolates were clearly separated from other Gaeumannomyces spp. Primers AV1 and AV3 amplified 1.3-kb portions of an avenacinase-like gene in the Phialophora isolates. Phylogenetic trees of the avenacinase-like gene in the Phialophora spp. also clearly separated them from other Gaeumannomyces spp. The Phialophora isolates were moderately virulent on wheat and barley and produced confined black lesions on the roots of wild oat and two oat cultivars. Among isolates tested for their sensitivity to 2,4-diacetylphloroglucinol (2,4-DAPG), the 90% effective dose values were 11.9 to 48.2 microg ml(-1). A representative Phialophora isolate reduced the severity of take-all on wheat caused by two different isolates of Gaeumannomyces graminis var. tritici. To our knowledge, this study provides the first report of an avenacinase-like gene in Phialophora spp. and demonstrated that the fungus is significantly less sensitive to 2,4-DAPG than G. graminis var. tritici.