Abstract
Pseudomonas fluorescens F113 is an effective biocontrol agent against Pythium ultimum, the causative agent of damping-off of sugarbeet seedlings. Biocontrol is mediated via the production of the anti-fungal metabolite 2,4-diacetylphloroglucinol (Phl). A genetic approach was used to further enhance the biocontrol ability of Fl 13. Two genetically modified (GM) strains, P. fluorescens F113Rif (pCU8.3) and P. fluorescens F113Rif (pCUP9), were developed for enhanced Ph1 production and assessed for biocontrol efficacy and impact on sugarbeet in microcosm experiments. The multicopy plasmid pCU8.3 contains the biosynthetic genes (phlA, C, B and D) and the putative permease gene (phlE) of F113 cloned into the rhizosphere stable plasmid pME6010, independent of external promoters. The plasmid pCUP9 consists of the Ph1 biosynthetic genes cloned downstream of the constitutive Plac promoter in pBBR1MCS. Introduction of pCU8.3 and pCUP9 into P. fluorescens F113 significantly altered the kinetics of Ph1 biosynthesis when grown in SA medium. A significant and substantial increase in Ph1 production by the GM strains was observed in the early logarithmic phase and stationary phase of growth compared with the wild-type strain. In microcosm, the two Ph1 overproducing strains proved to be as effective at controlling damping-off disease as the proprietary fungicide treatment, indicating the potential of genetic modification for plant disease control.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Preview
Unable to display preview. Download preview PDF.
References
Adams C and O’Gara F 1999 Molecular genetics of the interactions of Psedomonas with its diverse environments. Pseudomonas’ 99: Biotechnology and Pathology, Maui, Hawaii. 1-5 September 1999.
Agrios G N 1997 Plant Pathology. 4th edition. Academic Press, San Diego, CA.
Bakker P W and Schippers B 1987 Microbial cyanide production in the rhizosphere in relation to potato yield reduction and Pseudomonas spp.-mediated plant growth-stimulation. Soil Biol. Biochem. 19,451–470.
Bangera M G and Thomashow L S 1996 Characterisation of a genomic locus required for synthesis of the antibiotic 2,4-diacetylphloroglucinol by the biological control agent Pseudomonas fluorescens Q2-87. Mol. Plant-Microbe Interact. 9, 83–90.
Bull C T, Weiler, D M and Thomashow, L S 1991 Relationship between root colonisation and suppression of Gaeumannomyces graminis var. tritici by Pseudomonas fluorescens strain 2-79. Phytopathology 81, 954–959.
Campbell R 1994 Biological control of soil-borne diseases: some present problems and different approaches. Crop Protect. 13, 4–13.
Carroll H, Moënne-Loccoz Y, Dowling D N and O’Gara F 1995 Mutational disruption of the biosynthesis genes coding for the antifungal metabolite 2,4-diacetylphloroglucinol does not influence the ecological fitness of Pseudomonas fluorescens F113 in the rhizosphere of sugarbeets. Appl. Environ. Microbiol. 61, 3002–3007.
Cook R J 1993 Making greater use of introduced microorganisms for biological control of plant pathogens. Annu. Rev. Phytopathol. 31,53–80.
Delany I, Sheehan M M, Fenton A, Bardin S, Aarons S, and O’Gara F 2000 Regulation of production of the antifungal metabolite 2,4-diacetylphloroglucinol in Pseudomonas fluorescens F113: genetic analysis of phlF as a transcriptional repressor. Microbiology 146,537–546.
Dhingra O D and Sinclair J B 1986 Basic Plant Pathology Methods. CRC Press, Boca Raton, FL.
Ditta G, Stanfield S, Corbin D and Helsinki D 1980 Broad host range DNA cloning system for gram-negative bacteria: construction of a gene bank of Rhizobium meliloti. Proc. Natl. Acad. Sci. USA 77, 7347–7351.
Dowling D N and O’Gara F 1994 Metabolites of Pseudomonas involved in the biocontrol of plant diseases. Trends Biotechnol. 12, 133–141.
Dunne C, Crowley, J J, Moënne-Loccoz Y, Dowling D N, de Bruijn F J, and O’Gara F 1997 Biological control of Pythium ultimum by Stenotrophomonas maltophilia W81 is mediated by an extracellular proteolytic activity. Microbiology 143, 3921–3931.
Dunne C, Moënne-Loccoz Y, McCarthy J, Higgins P, Powell J, Dowling D N and O’Gara F 1997 Combining proteolytic and phloroglucinol-producing bacteria for improved biocontrol of Pythium-medisited damping-off of sugarbeet. Plant Pathol. 47, 299–307.
Farinha M A and Kropinski A M 1990 High efficiency electroporation of Pseudomonas aeruginosa using frozen cell suspensions. FEMS Microbiol. Lett. 70, 221–226.
Fenton A M, Stephens P M, Crowley J, O’Callaghan M and O’Gara F 1992 Exploitation of gene(s) involved in 2,4-diacetylphloroglucinol biosynthesis to confer a new biocontrol capability to a Pseudomonas strain. Appl. Environ. Microbiol. 58, 3873–3878.
Figurski D H and Helinski D R 1979 Replication of an origin-containing derivative of plasmid RK2 dependent on a plasmid function provided in trans. Proc. Natl. Acad. Sci. USA 76, 1648–1652.
Gould W D, Hagedorn C, Bardinelli T R and Zablotowicz R M 1985 New selective media for enumeration and recovery of fluorescent pseudomonads from various habitats. Appl. Environ. Microbiol. 49, 28–32.
Heeb S, Itoh Y, Nishijyo T, Schnider U, Keel C, Wade J, Walsh U, O’Gara F and Haas D 2000 Small, stable shuttle vectors based on the minimal pVSl replicon for use in Gram-negative plant-associated bacteria. Mol. Plant-Microbe Interact. 13, 232–237.
Hill D S, Stein J I, Torkewitz N R, Morse A M, Howell C R, Pach-latko J P, Becker J O and Ligon J M 1994 Cloning of genes involved in the synthesis of pyrrolnitrin from Pseudomonas fluorescens and role of pyrrolnitrin synthesis in biological control of plant disease. Appl. Environ. Microbiol. 60, 78–85.
Homma Y, Sato Z, Hirayama F, Konno K, Shirahama H and Suzui T 1989 Production of antibiotics by Pseudomonas cepacia as an agent for biological control of soilborne pathogens. Soil Biol. Biochem. 21,723–728.
Keel C, Schnider U, Maurhofer M, Voisard C, Laville J, Burger U, Wirthner P, Haas D and Défago G 1992 Suppression of root diseases by Pseudomonas fluorescens CGA0: importance of the bacterial secondary metabolite 2,4-diacetylphloroglucinol. Mol. Plant-Microbe Interact. 5, 4–13.
Keel C and Défago G 1997 Interactions between beneficial soil bacteria and root pathogens: mechanisms and ecological impact. In Multitrophic Interactions in Terrestrial Systems. Ed. A Gange and V K Brown, p 27–46. Blackwell Scientific, London.
Kovach M E, Phillips R W, Elzer P H, Roop II P H and Peterson K M 1994 pBBRIMCS: a broad-host-range cloning vector. Biotechniques 16, 800–802.
Leach L D 1986 Seedling diseases. In Compendium of Beet Diseases and Insects. Ed. E D Whitney and J E Duffus. The American Phytopathological Society, Minnesota, USA.
Lemanceau P, Bakker P A H M, de Kogel W J, Alabouvette C and Schippers B 1992 Effect of pseudobactin 358 production by Pseudomonas putida WCS358 on suppression of Fusarium wilt of carnations by nonpathogenic Fusarium oxysporum Fo47 Appl. Environ. Microbiol. 58, 2978–2982.
Levy E, Gough F J, Berlin K D, Guiana P W and Smith J T 1992 Inhibition of Septoria tritici and other phytopathogenic fungi and bacteria by Pseudomonas fluorescens and its antibiotics. Plant Pathol. 41,335–341.
Maurhofer M, Keel C, Haas D and Défago G 1995 Influence of plant species on disease suppression by Pseudomonas fluorescens strain CHA0 with enhanced antibiotic production. Plant Pathol. 44, 40–50.
Moënne-Loccoz Y, Powell J, Higgins P, McCarthy J and O’Gara F 1998 An investigation of the impact of biocontrol Pseudomonas fluorescens F113 on the growth of sugarbeet and the performance of subsequent clover-Rhizobium symbiosis. Appl. Soil Ecol. 7, 225–237.
Raaijmakers J M, Weiler D M and Thomashow L S 1997 Frequency of antibiotic-producing Pseudomonas spp. in natural environments. Appl. Environ. Microbiol. 63, 881–887.
Sambrook J, Frisch E F and Maniatis T 1989 Molecular Cloning: A Laboratory Manual. 2nd Edn. Cold Spring Harbour Laboratory Press, Cold Spring Harbour, NY.
Scher F M and Baker R 1982 Effect of Pseudomonas putida and a synthetic iron chelator on induction of soil suppressiveness to Fusarium wilt pathogens. Phytopathology 72, 1567–1573.
Schnider-Keel U, Seematter A, Maurhofer M, Blumer C, Duffy B, Gigot-Bonnefoy C, Reimmann C, Notz R, Défago G, Haas D and Keel C 2000 Autoinduction of 2,4-diacetylphloroglucinol biosynthesis in the biocontrol agent Pseudomonas fluorescens CHAO and repression by the bacterial metabolites salicylate and pyoluteorin. J. Bacteriol. 182, 1215–1225.
Shanahan P, O’Sullivan D J, Simpson P, Glennon J D and O’Gara F 1992 Isolation of 2,4-diacetylphloroglucinol from a fluorescent pseudomonad and investigation of physiological parameters influencing its production. Appl. Environ. Microbiol. 58, 353–358.
Thomashow L S, Bangera M G, Bonsall R F, Kim R F, Raaijmakers J and Weiler D M 1997 2,4-Diacetylphloroglucinol, a key antibiotic in soilborne pathogen suppression be fluorescent Pseudomonas spp. In Biology of Plant-Microbe Interactions. Ed. G Stacey, B Mullin and P M Gresshoff. Proceedings of the 8th International Symposium on Molecular Plant-Microbe Interactions. Knoxville, TN, USA.
Thomashow L S and Weiler D M 1988 Role of a phenazine antibiotic from Pseudomonas fluorescens in biological control of Gaeumannomyces graminis var. tritici. J. Bacteriol. 170, 3499–3508.
Vincent M N, Harrison J M, Brackin J M, Kovacevich, Mukerji P, Weiler D M and Pierson E A 1991 Genetic analysis of the antifungal activity of a soilborne Pseudomonas aureofaciens strain. Appl. Environ. Microbiol. 57, 2928–2934.
Voisard C, Keel C, Haas D and Défago G 1989 Cyanide production by Pseudomonas fluorescens helps suppress black root rot of tobacco under gnotobiotic conditions. EMBO J. 8, 351–358.
Author information
Authors and Affiliations
Editor information
Rights and permissions
Copyright information
© 2002 Springer Science+Business Media Dordrecht
About this chapter
Cite this chapter
Delany, I.R., Walsh, U.F., Ross, I., Fenton, A.M., Corkery, D.M., O’Gara, F. (2002). Enhancing the biocontrol efficacy of Pseudomonas fluorescens F113 by altering the regulation and production of 2,4-diacetylphloroglucinol. In: Powlson, D.S., Bateman, G.L., Davies, K.G., Gaunt, J.L., Hirsch, P.R. (eds) Interactions in the Root Environment: An Integrated Approach. Developments in Plant and Soil Sciences, vol 96. Springer, Dordrecht. https://doi.org/10.1007/978-94-010-0566-1_19
Download citation
DOI: https://doi.org/10.1007/978-94-010-0566-1_19
Publisher Name: Springer, Dordrecht
Print ISBN: 978-94-010-3925-3
Online ISBN: 978-94-010-0566-1
eBook Packages: Springer Book Archive