Characterization of CMR5c and CMR12a, novel fluorescent Pseudomonas strains from the cocoyam rhizosphere with biocontrol activity

Aim: To screen for novel antagonistic Pseudomonas strains producing both phenazines and biosurfactants that are as effective as Pseudomonas aeruginosa PNA1 in the biocontrol of cocoyam root rot caused by Pythium myriotylum. Material and Results: Forty pseudomonads were isolated from the rhizosphere of healthy white and red cocoyam plants appearing in natural, heavily infested fields in Cameroon. In vitro tests demonstrated that Py. myriotylum antagonists could be retrieved from the red cocoyam rhizosphere. Except for one isolate, all antagonistic isolates produced phenazines. Results from whole-cell protein profiling showed that the antagonistic isolates are different from other isolated pseudomonads, while BOX-PCR revealed high genomic similarity among them. 16S rDNA sequencing of two representative strains within this group of antagonists confirmed their relatively low similarity with validly described Pseudomonas species. These antagonists are thus provisionally labelled as unidentified Pseudomonas strains. Among the antagonists, Pseudomonas CMR5c and CMR12a were selected because of their combined production of phenazines and biosurfactants. For strain CMR5c also, production of pyrrolnitrin and pyoluteorin was demonstrated. Both CMR5c and CMR12a showed excellent in vivo biocontrol activity against Py. myriotylum to a similar level as Ps. aeruginosa PNA1. Conclusion: Pseudomonas CMR5c and CMR12a were identified as novel and promising biocontrol agents of Py. myriotylum on cocoyam, producing an arsenal of antagonistic metabolites. Significance and Impact of the Study: Present study reports the identification of two newly isolated fluorescent Pseudomonas strains that can replace the opportunistic human pathogen Ps. aeruginosa PNA1 in the biocontrol of cocoyam root rot and could be taken into account for the suppression of many plant pathogens

Impact of wild-type and genetically modified Pseudomonas fluorescens on soil enzyme activities and microbial population structure in the rhizosphere of pea

The definitive version is available at www.blackwell-synergy.com. Copyright Blackwell Publishing DOI : 10.1046/j.1365-294x.1998.00367.xThe aim of this work was to determine the impact of wild type along with functionally and non-functionally modified Pseudomonas fluorescens strains in the rhizosphere. The wild type F113 strain carried a gene encoding the production of the antibiotic 2,4 diacetylphloroglucinol (DAPG) useful in plant disease control, and was marked with a lacZY gene cassette. The first modified strain was a functional modification of strain F113 with repressed production of DAPG, creating the DAPG negative strain F113 G22. The second paired comparison was a non-functional modification of wild type (unmarked) strain SBW25, constructed to carry marker genes only, creating strain SBW25 EeZY-6KX. Significant perturbations were found in the indigenous bacterial population structure, with the F113, (DAPG+) strain causing a shift towards slower growing colonies (K strategists) compared with the non-antibiotic producing derivative (F113 G22) and the SBW25 strains. The DAPG+ strain also significantly reduced, in comparison with the other inocula, the total Pseudomonas populations but did not affect the total microbial populations. The survival of F113 and F113 G22 were an order of magnitude lower than the SBW 25 strains. The DAPG+ strain caused a significant decrease in the shoot to root ratio in comparison to the control and other inoculants, indicating plant stress. F113 increased soil alkaline phosphatase, phosphodiesterase and aryl sulphatase activities compared to the other inocula, which themselves reduced the same enzyme activities compared to the control. In contrast to this, the -glucosidase, -galactosidase and N-acetyl glucosaminidase activities decreased with the inoculation of the DAPG+ strain. These results indicate that soil enzymes are sensitive to the impact of GMM inoculation.Peer reviewe

Evolutionary patchwork of an insecticidal toxin shared between plant-associated pseudomonads and the insect pathogens Photorhabdus and Xenorhabdus

Background: Root-colonizing fluorescent pseudomonads are known for their excellent abilities to protect plants against soil-borne fungal pathogens. Some of these bacteria produce an insecticidal toxin (Fit) suggesting that they may exploit insect hosts as a secondary niche. However, the ecological relevance of insect toxicity and the mechanisms driving the evolution of toxin production remain puzzling. Results: Screening a large collection of plant-associated pseudomonads for insecticidal activity and presence of the Fit toxin revealed that Fit is highly indicative of insecticidal activity and predicts that Pseudomonas protegens and P. chlororaphis are exclusive Fit producers. A comparative evolutionary analysis of Fit toxin-producing Pseudomonas including the insect-pathogenic bacteria Photorhabdus and Xenorhadus, which produce the Fit related Mcf toxin, showed that fit genes are part of a dynamic genomic region with substantial presence/absence polymorphism and local variation in GC base composition. The patchy distribution and phylogenetic incongruence of fit genes indicate that the Fit cluster evolved via horizontal transfer, followed by functional integration of vertically transmitted genes, generating a unique Pseudomonas-specific insect toxin cluster. Conclusions: Our findings suggest that multiple independent evolutionary events led to formation of at least three versions of the Mcf/Fit toxin highlighting the dynamic nature of insect toxin evolution

The rhizosphere: a playground and battlefield for soilborne pathogens and beneficial microorganisms

The rhizosphere is a hot spot of microbial interactions as exudates released by plant roots are a main food source for microorganisms and a driving force of their population density and activities. The rhizosphere harbors many organisms that have a neutral effect on the plant, but also attracts organisms that exert deleterious or beneficial effects on the plant. Microorganisms that adversely affect plant growth and health are the pathogenic fungi, oomycetes, bacteria and nematodes. Most of the soilborne pathogens are adapted to grow and survive in the bulk soil, but the rhizosphere is the playground and infection court where the pathogen establishes a parasitic relationship with the plant. The rhizosphere is also a battlefield where the complex rhizosphere community, both microflora and microfauna, interact with pathogens and influence the outcome of pathogen infection. A wide range of microorganisms are beneficial to the plant and include nitrogen-fixing bacteria, endo- and ectomycorrhizal fungi, and plant growth-promoting bacteria and fungi. This review focuses on the population dynamics and activity of soilborne pathogens and beneficial microorganisms. Specific attention is given to mechanisms involved in the tripartite interactions between beneficial microorganisms, pathogens and the plant. We also discuss how agricultural practices affect pathogen and antagonist populations and how these practices can be adopted to promote plant growth and health

Isolation, cloning and characterization of phlB gene from an Indian strain of Gram negative soil bacteria Pseudomonas fluorescens

412-419Phloroglucinol (PHL) or 2,4-diacetylphloroglucinol (DAPG) is a polyketide compound that exhibits broad spectrum anti-bacterial and antifungal properties against soil-borne plant pathogens. It is produced by gram negative bacterium Pseudomonas (fluorescent Pseudomonad) which renders them a potential biological control agent. The process of PHL production is accomplished by a well coordinated activity of four (phlABCD) genes arranged in an operon. All four genes are essential and indispensable for biosynthesis of DAPG from precursor malonyl CoA. Here, we report cloning and analysis of phlB gene from an Indian strain of Pseudomonas fluorescens. Further, we predicted the structure of PHLB protein and analyzed its intrinsic details. Stereo-chemical properties were analyzed by Ramachandran plot and secondary structure was retrieved by PDB sum. Our study provides insight into the structure of PHLB protein and revealed that though Pseudomonas PHLB lacks conserved motifs and domains it is indispensable for monoacetyl phloroglucinol (MAPG) synthesis

Wheat Cultivar-Specific Selection of 2,4-Diacetylphloroglucinol-Producing Fluorescent \u3ci\u3ePseudomonas\u3c/i\u3e Species from Resident Soil Populations

An emerging body of evidence indicates a role for plant genotype as a determinant of the species and genetic composition of the saprophytic microbial community resident to the rhizosphere. In this study, experiments were conducted to determine the capacity of five different wheat cultivars to enhance resident populations and support introduced strains of 2,4-diacetylphloroglucinol (2,4-DAPG)-producing fluorescent pseudomonads, a group of bacteria known to provide biological control of several soilborne diseases. When soils were cropped with three successive 28-day growth cycles of wheat, the 2,4- DAPG-producing strains were consistently recovered from the rhizosphere of the cultivar Lewjain, and commonly were present at populations higher than those recovered from other wheat cultivars. Based on restriction fragment length polymorphism and sequence analyses of phlD, a key gene involved in 2,4-DAPG production, two previously undefined phlD+ genotypes, referred to as genotypes PfZ and PfY, were discovered. Wheat cultivar Lewjain was the primary source of genotype PfY while cultivar Penawawa yielded the majority of genotype PfZ. Based on 16S rDNA sequence analysis, both new phlD genotypes were classified as P. fluorescens. Comparison of the rhizosphere competence of 2,4-DAPG-producing P. fluorescens Q2-87 (genotype B) and P. fluorescens LR3-A28 (genotype PfY) showed that both strains persisted at similar populations in the rhizosphere of all cultivars tested over a 30 day period when introduced as a seed inoculant. However, when strain LR3-A28 was applied as a soil inoculant, this strain was recovered at higher populations from the rhizosphere of wheat cultivar Lewjain than from the rhizospheres of two other cultivars. No cultivar effects were shown for strain Q2-87. Collectively, these results add further to evidence indicating a degree of specificity in interactions between plant cultivars and specific members of the saprophytic microbial community. Furthermore, as 2,4- DAPG-producing fluorescent Pseudomonas spp. have a central role in the spontaneous reduction in severity of take-all disease of wheat in response to continuous wheat monoculture, we postulate that the use of specific cultivars, such as Lewjain, which possess a superior capacity to enhance resident soil populations of these bacteria may have potential to reduce the length of the monoculture period required to induce natural suppressiveness of soils toward this disease

Draft Genome Sequence of Pseudomonas sp. Strain LD120, Isolated from the Marine Alga Saccharina latissima

We report the draft genome sequence of Pseudomonas sp. strain LD120, which was isolated from a brown macroalga in the Baltic Sea. The genome of this marine Pseudomonas protegens subgroup bacterium harbors biosynthetic gene clusters for toxic metabolites typically produced by members of this Pseudomonas subgroup, including 2,4-diacetylphloroglucinol, pyoluteorin, and rhizoxin analogs.ISSN:2576-098

Prevalence of fluorescent pseudomonads producing antifungal phloroglucinols and/or hydrogen cyanide in soils naturally suppressive or conducive to tobacco black root rot

Certain soils from Morens, Switzerland, are naturally suppressive to Thielaviopsis basicola-mediated black root rot of tobacco, and fluorescent pseudomonads are involved in this suppressiveness. Here, we compared two conducive, one moderately suppressive and one suppressive soil from Morens. Disease levels on tobacco after heavy T. basicola inoculation varied from 29% to 85% for the two conducive soils, 10% to 78% for the moderately suppressive soil and 11% to 42% for the suppressive soil, depending on time of the year. In the absence of T. basicola inoculation, disease levels were between 0% and 40% and varied also in time. Fluorescent pseudomonads were isolated from the rhizosphere and roots of tobacco subjected to T. basicola inoculation and characterized for production of the biocontrol metabolites 2,4-diacetylphloroglucinol (Phl) and HCN. No difference in population size was found between the suppressive and the conducive soils for total, Phl+ and HCN+ fluorescent pseudomonads colonizing the rhizosphere or roots of tobacco. Yet, the percentage of Phl+ isolates was significantly higher (30-32% vs. 6-11%) in the rhizosphere and roots for plants grown in the suppressive soil compared with the moderately suppressive and conducive soils. Different restriction profiles for phlD, one of the Phl biosynthetic genes, were often found when analyzing Phl+ isolates colonizing the same plant. Most phlD alleles were recovered from both suppressive and conducive soils, except one allele found only in root isolates from the suppressive soi