Geneviève Défago

Publications (47)168.93 Total impact

• Source

  Available from: Vincent Walker

  Article: Comparison of prominent Azospirillum strains in Azospirillum-Pseudomonas-Glomus consortia for promotion of maize growth.

  Olivier Couillerot, Augusto Ramírez-Trujillo, Vincent Walker, Andreas von Felten, Jan Jansa, Monika Maurhofer, Geneviève Défago, Claire Prigent-Combaret, Gilles Comte, Jesus Caballero-Mellado, Yvan Moënne-Loccoz
  [show abstract] [hide abstract]
  ABSTRACT: Azospirillum are prominent plant growth-promoting rhizobacteria (PGPR) extensively used as phytostimulatory crop inoculants, but only few studies are dealing with Azospirillum-containing mixed inocula involving more than two microorganisms. We compared here three prominent Azospirillum strains as part of three-component consortia including also the PGPR Pseudomonas fluorescens F113 and a mycorrhizal inoculant mix composed of three Glomus strains. Inoculant colonization of maize was assessed by quantitative PCR, transcription of auxin synthesis gene ipdC (involved in phytostimulation) in Azospirillum by RT-PCR, and effects on maize by secondary metabolic profiling and shoot biomass measurements. Results showed that phytostimulation by all the three-component consortia was comparable, despite contrasted survival of the Azospirillum strains and different secondary metabolic responses of maize to inoculation. Unexpectedly, the presence of Azospirillum in the inoculum resulted in lower phytostimulation in comparison with the Pseudomonas-Glomus two-component consortium, but this effect was transient. Azospirillum's ipdC gene was transcribed in all treatments, especially with three-component consortia, but not with all plants and samplings. Inoculation had no negative impact on the prevalence of mycorrhizal taxa in roots. In conclusion, this study brought new insights in the functioning of microbial consortia and showed that Azospirillum-Pseudomonas-Glomus three-component inoculants may be useful in environmental biotechnology for maize growth promotion.
  Applied Microbiology and Biotechnology 07/2012; · 3.42 Impact Factor
• Article: Relationship between aggressiveness of Stagonospora sp. isolates on field and hedge bindweeds, and in vitro production of fungal metabolites cercosporin, elsinochrome A and leptosphaerodione

  Monday O. Ahonsi, Monika Maurhofer, Désirée Boss, Geneviève Défago
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  ABSTRACT: Stagonospora convolvuli LA39, an effective biocontrol agent of Convolvulus arvensis (field bindweed) and Calystegia sepium (hedge bindweed) produces phytotoxic metabolites leptosphaerodione and elsinochrome A. Stagonospora isolate 214Caa produces the toxin cercosporin. If toxic metabolite production is not linked to the pathogenic ability of the fungus on bindweeds, selection of aggressive strains with limited or no production of the metabolites would reduce any perceived risk of using strains of the fungus as a mycoherbicide. Therefore, 30 isolates of Stagonospora sp. including LA39 and 214Caa were characterised for aggressiveness on both bindweeds, and production of the three metabolites. Nine isolates were more aggressive than LA39 on both bindweeds. Classification of isolates based on metabolite type agreed largely with previous similar characterisation based on polymerase chain reaction-restriction fragment length polymorphism of internal transcribed spacer of ribosomal DNA. Cercosporin producers produced neither leptosphaerodione nor elsinochrome A and together with isolates that produce none of the three metabolites, were less pathogenic on bindweeds. Conversely, there was a positive correlation between elsinochrome A and leptosphaerodione production, and each was positively correlated with aggressiveness of isolates on both bindweeds. Generally, any isolate where elsinochrome A was not detected was not aggressive on any of the two bindweeds. This probably implies that selecting elsinochrome A-negative, but aggressive Stagonospora strain(s) may be difficult. However, aggressive isolates may not produce elsinochrome A in planta at levels that could constitute any risk in the environment. In a preliminary attempt to determine the levels of elsinochrome A and leptosphaerodione produced in diseased bindweeds, none of the toxins was detected in Stagonospora infected bindweed leaves. Detailed investigation focusing on the detection and quantification of in planta production of elsinochrome A by Stagonospora isolates, and determination of the fate of elsinochrome A in the environment, and its relationship with leptosphaerodione may be essential. Similarly, development of molecular tools to monitor the mycoherbicide following field application is vital.
  European Journal of Plant Pathology 04/2012; 111(3):203-215. · 1.41 Impact Factor
• Article: Evolutionary history of synthesis pathway genes for phloroglucinol and cyanide antimicrobials in plant-associated fluorescent pseudomonads.

  Michele Frapolli, Joël F Pothier, Geneviève Défago, Yvan Moënne-Loccoz
  [show abstract] [hide abstract]
  ABSTRACT: Plant-beneficial fluorescent Pseudomonas spp. play important ecological roles. Here, their evolutionary history was investigated by a multilocus approach targeting genes involved in synthesis of secondary antimicrobial metabolites implicated in biocontrol of phytopathogens. Some of these genes were proposed to be ancestral, and this was investigated using a worldwide collection of 30 plant-colonizing fluorescent pseudomonads, based on phylogenetic analysis of 14 loci involved in production of 2,4-diacetylphloroglucinol (phlACBDE, phlF, intergenic locus phlA/phlF), hydrogen cyanide (hcnABC, anr) or global regulation of secondary metabolism (gacA, gacS, rsmZ). The 10 housekeeping loci rrs, dsbA, gyrB, rpoD, fdxA, recA, rpoB, rpsL, rpsG, and fusA served as controls. Each strain was readily distinguished from the others when considering allelic combinations for these 14 biocontrol-relevant loci. Topology comparisons based on Shimodaira-Hasegawa tests showed extensive incongruence when comparing single-locus phylogenetic trees with one another, but less when comparing (after sequence concatenation) trees inferred for genes involved in 2,4-diacetylphloroglucinol synthesis, hydrogen cyanide synthesis, or secondary metabolism global regulation with trees for housekeeping genes. The 14 loci displayed linkage disequilibrium, as housekeeping loci did, and all 12 protein-coding loci were subjected to purifying selection except for one positively-selected site in HcnA. Overall, the evolutionary history of Pseudomonas genes involved in synthesis of secondary antimicrobial metabolites important for biocontrol functions is in fact similar to that of housekeeping genes, and results suggest that they are ancestral in pseudomonads producing hydrogen cyanide and 2,4-diacetylphloroglucinol.
  Molecular Phylogenetics and Evolution 03/2012; 63(3):877-90. · 3.61 Impact Factor
• Article: Pseudomonas protegens sp. nov., widespread plant-protecting bacteria producing the biocontrol compounds 2,4-diacetylphloroglucinol and pyoluteorin.

  Alban Ramette, Michele Frapolli, Marion Fischer-Le Saux, C Gruffaz, Jean-Marie Meyer, Geneviève Défago, Laurent Sutra, Yvan Moënne-Loccoz
  [show abstract] [hide abstract]
  ABSTRACT: Fluorescent Pseudomonas strains producing the antimicrobial secondary metabolite 2,4-diacetylphloroglucinol (Phl) play a prominent role in the biocontrol of plant diseases. A subset of Phl-producing fluorescent Pseudomonas strains, which can additionally synthesize the antimicrobial compound pyoluteorin (Plt), appears to cluster separately from other fluorescent Pseudomonas spp. based on 16S rRNA gene analysis and shares at most 98.4% 16S rRNA gene sequence identity with any other Pseudomonas species. In this study, a polyphasic approach based on molecular and phenotypic methods was used to clarify the taxonomy of representative Phl(+) Plt(+) strains isolated from tobacco, cotton or wheat on different continents. Phl(+) Plt(+) strains clustered separately from their nearest phylogenetic neighbors (i.e. species from the 'P. syringae', 'P. fluorescens' and 'P. chlororaphis' species complexes) based on rpoB, rpoD or gyrB phylogenies. DNA-DNA hybridization experiments clarified that Phl(+) Plt(+) strains formed a tight genomospecies that was distinct from P. syringae, P. fluorescens, or P. chlororaphis type strains. Within Phl(+) strains, the Phl(+) Plt(+) strains were differentiated from other biocontrol fluorescent Pseudomonas strains that produced Phl but not Plt, based on phenotypic and molecular data. Discriminative phenotypic characters were also identified by numerical taxonomic analysis and siderotyping. Altogether, this polyphasic approach supported the conclusion that Phl(+) Plt(+) fluorescent Pseudomonas strains belonged to a novel species for which the name Pseudomonas protegens is proposed, with CHA0(T) (=CFBP 6595(T), =DSM 19095(T)) as the type strain.
  Systematic and Applied Microbiology 03/2011; 34(3):180-8. · 3.37 Impact Factor
• Article: Novel T-RFLP method to investigate six main groups of 2,4-diacetylphloroglucinol-producing pseudomonads in environmental samples.

  Andreas von Felten, Joana Beatrice Meyer, Geneviève Défago, Monika Maurhofer
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  ABSTRACT: Strains of fluorescent pseudomonads producing 2,4-diacetylphloroglucinol (DAPG) are involved in the protection of plant roots against soil-borne plant pathogens. Recently, a multilocus sequence analysis of a world wide collection of DAPG-producers led to the identification of six main groups (A-F). In this study a T-RFLP method based on the phlD gene was developed to efficiently identify the members of these six groups in environmental samples. A combination of six restriction enzymes was identified which leads to group specific terminal fragments (T-RF). The detection limit of the phlD-T-RFLP method was determined for the two P. fluorescens strains F113 (group B) and CHA0 (group F) in rhizosphere samples and was found to be 5×10(3)CFU/g and 5×10(4)CFU/g respectively. PhlD-T-RFLP and phlD-DGGE analysis of wheat and maize root samples from greenhouse and field revealed similarly the presence of multilocus groups A, B and D. However, they were more frequently detected with phlD-T-RFLP. Additionally, groups C and F were detected in greenhouse samples but only by phlD-T-RFLP and not by phlD-DGGE. In conclusion, the new phlD-T-RFLP method proved to be a fast and reliable method to detect strains of the six main groups of DAPG-producers in environmental samples with an improved detection limit compared to phlD-DGGE.
  Journal of microbiological methods 12/2010; 84(3):379-87. · 2.43 Impact Factor
• Source

  Available from: Cesare Gessler

  Article: Selection of a biocontrol agent based on a potential mechanism of action: degradation of nicotinic acid, a growth factor essential for Erwinia amylovora.

  Thomas Paternoster, Geneviève Défago, Brion Duffy, Cesare Gessler, Ilaria Pertot
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  ABSTRACT: This work describes a medium-based screening method for selecting microbial biocontrol agents against Erwinia amylovora based on the degradation of a specific growth factor. Erwinia amylovora, the causal agent of the devastating fire blight disease, requires nicotinic acid or nicotinamide as an essential growth factor. Potential biocontrol agents are either selected for antimicrobial production in plate or directly on immature pears or apple blossoms. In this work, we have attempted to streamline the selection of a new potential biocontrol agent with a lower risk of non-target effects by isolation based on the ability to degrade nicotinic acid in vitro, using therefore few plant materials. A total of 735 bacteria and 1237 yeast were isolated from apple blossoms and pre-screened for nicotinic acid-degradation. Pseudomonas rhizosphaerae strain JAN was able to degrade both nicotinic acid and nicotinamide. Mutants deficient in this ability were constructed. JAN, but not the mutants, controlled E. amylovora on pear slices. On detached apple blossoms, JAN colonized apple hypanthia and strongly suppressed E. amylovora growth. Under greenhouse conditions, JAN was more effective in controlling blossom blight than P. fluorescens A506, a commercial biocontrol agent of fire blight unable to degrade nicotinic acid and nicotinamide.
  International Microbiology 12/2010; 13(4):195-206. · 1.80 Impact Factor
• Article: Interplay between wheat cultivars, biocontrol pseudomonads, and soil.

  Joana Beatrice Meyer, Matthias Peter Lutz, Michele Frapolli, Maria Péchy-Tarr, Laurène Rochat, Christoph Keel, Geneviève Défago, Monika Maurhofer
  [show abstract] [hide abstract]
  ABSTRACT: There is a significant potential to improve the plant-beneficial effects of root-colonizing pseudomonads by breeding wheat genotypes with a greater capacity to sustain interactions with these bacteria. However, the interaction between pseudomonads and crop plants at the cultivar level, as well as the conditions which favor the accumulation of beneficial microorganisms in the wheat rhizosphere, is largely unknown. Therefore, we characterized the three Swiss winter wheat (Triticum aestivum) cultivars Arina, Zinal, and Cimetta for their ability to accumulate naturally occurring plant-beneficial pseudomonads in the rhizosphere. Cultivar performance was measured also by the ability to select for specific genotypes of 2,4-diacetylphloroglucinol (DAPG) producers in two different soils. Cultivar-specific differences were found; however, these were strongly influenced by the soil type. Denaturing gradient gel electrophoresis (DGGE) analysis of fragments of the DAPG biosynthetic gene phlD amplified from natural Pseudomonas rhizosphere populations revealed that phlD diversity substantially varied between the two soils and that there was a cultivar-specific accumulation of certain phlD genotypes in one soil but not in the other. Furthermore, the three cultivars were tested for their ability to benefit from Pseudomonas inoculants. Interestingly, Arina, which was best protected against Pythium ultimum infection by inoculation with Pseudomonas fluorescens biocontrol strain CHA0, was the cultivar which profited the least from the bacterial inoculant in terms of plant growth promotion in the absence of the pathogen. Knowledge gained of the interactions between wheat cultivars, beneficial pseudomonads, and soil types allows us to optimize cultivar-soil combinations for the promotion of growth through beneficial pseudomonads. Additionally, this information can be implemented by breeders into a new and unique breeding strategy for low-input and organic conditions.
  Applied and environmental microbiology 09/2010; 76(18):6196-204. · 3.69 Impact Factor
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  Available from: Miro Svercel

  Article: Distribution of Pseudomonas populations harboring phlD or hcnAB biocontrol genes is related to depth in vineyard soils

  Miroslav Svercel, Jérôme Hamelin, Brion Duffy, Yvan Moënne-Loccoz, Geneviève Défago
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  ABSTRACT: The abundance and population structure of pseudomonads in soils collected from long-(1006 years) and short-(54 years) term grapevine monocultures in Switzerland were examined across five soil horizons within the 1.20–1.35 m range. Soil samples were baited with grapevine, and rhizosphere pseudomonads containing the biocontrol genes phlD (2,4-diacetylphloroglucinol synthesis) and/or hcnAB (hydrogen cyanide synthesis) were analyzed by MPN-PCR. The numbers of total, phlD+ and hcnAB+ pseudomonads decreased with depth by 1.5–2 log (short-term monoculture) and 3–3.5 log (long-term monoculture). In addition, the percentages of phlD+ (except in short-term monoculture) and hcnAB+ pseudomonads were also lower in deeper horizons. RFLP-profiling of phlD+ and hcnAB+ pseudomonads revealed three phlD and twelve hcnAB alleles overall, but the number of alleles for both decreased in relation to depth. The only phlD allele found in deeper horizons was also found in topsoil, whereas one hcnAB allele (k) found in deeper horizons in long-term monoculture was absent in the topsoil. This suggests that certain Pseudomonas ecotypes are adapted to specific depths. Four hcnAB alleles enabled discrimination between monocultures. We conclude that soil depth is a factor selecting phlD and hcnAB genotypes, and that the allelic diversity of the two biocontrol genes decreases with depth.
  Soil Biology and Biochemistry 03/2010; 42(3):466-472. · 3.50 Impact Factor
• Article: Quantification of Pseudomonas fluorescens strains F113, CHA0 and Pf153 in the rhizosphere of maize by strain-specific real-time PCR unaffected by the variability of DNA extraction efficiency.

  Andreas Von Felten, Geneviève Défago, Monika Maurhofer
  [show abstract] [hide abstract]
  ABSTRACT: Pseudomonas fluorescens strains F113 and CHA0 are well-known plant growth-promoting rhizobacteria (PGPR) often used as model strains in biocontrol experiments. To monitor their persistence in large scale field experiments, culture-independent methods are needed. In this study, a strain-specific real-time PCR quantification tool was developed based on sequence-characterized amplified regions (SCAR) for P. fluorescens strains F113, CHA0 and Pf153. Differences in DNA extraction efficiencies from rhizosphere samples were circumvented using plasmid APA9 as internal standard to normalize C(T) values after real-time amplification. The detection limits of the real-time PCR assays for all three strains were approximately 10 cells for genomic DNA and 10(4)cells/g rhizosphere for maize samples grown in different natural soils. Population sizes of the three strains in the rhizosphere of maize measured by the new real-time PCR approaches were similar to those measured by most probable number (MPN)-PCR. A persistence study of the three strains indicated that the strains persisted differently over a period of 5weeks. In conclusion the newly developed real-time PCR approach is a fast and resource efficient method for monitoring individual biocontrol strains in natural soil, which makes it an apt quantification tool for future large-scale field experiments.
  Journal of microbiological methods 02/2010; 81(2):108-15. · 2.43 Impact Factor
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  Available from: Miro Svercel

  Article: Effect of long‐term vineyard monoculture on rhizosphere populations of pseudomonads carrying the antimicrobial biosynthetic genes phlD and/or hcnAB

  Miroslav Svercel, Danilo Christen, Yvan Moënne-Loccoz, Brion Duffy, Geneviève Défago
  [show abstract] [hide abstract]
  ABSTRACT: The impact of repeated culture of perennial plants (i.e. in long-term monoculture) on the ecology of plant-beneficial bacteria is unknown. Here, the influence of extremely long-term monocultures of grapevine (up to 1603 years) on rhizosphere populations of fluorescent pseudomonads carrying the biosynthetic genes phlD for 2,4-diacetylphloroglucinol and/or hcnAB for hydrogen cyanide was determined. Soils from long-term and adjacent short-term monoculture vineyards (or brushland) in four regions of Switzerland were baited with grapevine or tobacco plantlets, and rhizosphere pseudomonads were studied by most probable number (MPN)-PCR. Higher numbers and percentages of phlD+ and of hcnAB+ rhizosphere pseudomonads were detected on using soil from long-term vineyards. On focusing on phlD, restriction fragment length polymorphism profiling of the last phlD-positive MPN wells revealed seven phlD alleles (three exclusively on tobacco, thereof two new ones). Higher numbers of phlD alleles coincided with a lower prevalence of the allele displayed by the well-studied biocontrol strain Pseudomonas fluorescens F113. The prevalence of this allele was 35% for tobacco in long-term monoculture soils vs. >60% in the other three cases. We conclude that soils from long-term grapevine monocultures represent an untapped resource for isolating novel biocontrol Pseudomonas strains when tobacco is used as bait.
  FEMS Microbiology Ecology 03/2009; 68(1):25 - 36. · 3.41 Impact Factor
• Article: A new DGGE protocol targeting 2,4-diacetylphloroglucinol biosynthetic gene phlD from phylogenetically contrasted biocontrol pseudomonads for assessment of disease-suppressive soils.

  Michele Frapolli, Yvan Moënne-Loccoz, Joana Meyer, Geneviève Défago
  [show abstract] [hide abstract]
  ABSTRACT: In the rhizosphere, biocontrol pseudomonads producing 2,4-diacetylphloroglucinol (Phl) can protect plants from soil-borne pathogens. DGGE of phlD has been proposed to monitor these bacteria, but two distinct protocols were needed for analysis of both the 'Pseudomonas fluorescens' species complex and the strains from rrs restriction group ARDRA-1. Here, a single DGGE protocol performed on 668-bp GC-clamp-containing phlD amplicons was effective with both types of pseudomonads, and 36 reference biocontrol strains from the 'P. fluorescens' complex or group ARDRA-1 gave a total of 11 distinct DGGE bands. phlD amplicons with at least two to seven nucleotidic differences could be discriminated, and the discrimination level was similar to that of phlD restriction analysis with four enzymes. Multiple phlD-DGGE bands were obtained when studying rhizosphere soil containing indigenous phlD+ pseudomonads, and phlD diversity was higher when DGGE was implemented after incubation of tobacco rhizosphere extracts in semi-selective medium (MPN approach) in comparison with approaches based on direct analysis of rhizosphere DNA extracts or assessment of phlD+colonies. phlD-DGGE profiles differed for a soil suppressive and a soil conducive to black root rot of tobacco, and each soil yielded new phlD sequences. In conclusion, this DGGE protocol was useful for monitoring indigenous rhizosphere consortia of phlD+ pseudomonads.
  FEMS Microbiology Ecology 07/2008; 64(3):468-81. · 3.41 Impact Factor
• Chapter: Biocontrol of Root Diseases by Pseudomonas fluorescens CHA0: Current Concepts and Experimental Approaches

  Christophe Voisard, Carolee T. Bull, Christoph Keel, Jacques Laville, Monika Maurhofer, Ursula Schnider, Geneviève Défago, Dieter Haas
  12/2007: pages 67 - 89; , ISBN: 9783527615810
• Article: ISSR fingerprinting for the assessment of the bindweed biocontrol agent Stagonospora convolvuli LA39 after field release.

  Désirée Boss, Monika Maurhofer, Marcello Zala, Geneviève Défago, Patrick C Brunner
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  ABSTRACT: To develop a molecular identification method based on ISSR fingerprints to monitor the fungal leaf pathogen Stagonospora convolvuli LA39 used to biologically control bindweeds after a field release. The developed method proved to be suitable to clearly distinguish LA39 from resident Stagonospora spp. and was applied in two field experiments. First, the environmental persistence of LA39 was assessed in an overwintering experiment. LA39 could be re-isolated from infected bindweed 1 year after field application, but with very low frequency of occurrence. Secondly, LA39 was applied in an area with natural bindweed infestation and re-isolated from infected bindweed. The dispersal of LA39 during one season was poor (4-5 m). ISSR fingerprinting has been shown to be a valuable tool to monitor the environmental fate of S. convolvuli in the field. It is concluded that an LA39-based mycoherbicide will have minimal environmental impact caused by the restricted mobility, poor proliferation and poor persistence over seasons of LA39. Studies about the dispersal and survival of biocontrol agents after field release as well as the development of methods needed for this purpose are indispensable for a comprehensive risk assessment for biocontrol agents.
  Letters in Applied Microbiology 10/2007; 45(3):244-51. · 1.62 Impact Factor
• Article: Multilocus sequence analysis of biocontrol fluorescent Pseudomonas spp. producing the antifungal compound 2,4-diacetylphloroglucinol.

  Michele Frapolli, Geneviève Défago, Yvan Moënne-Loccoz
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  ABSTRACT: The genetic and evolutionary relationship among 2,4-diacetylphloroglucinol (Phl)-producing pseudomonads that protect plants from soil-borne pathogens were investigated by multilocus sequence typing. A total of 65 pseudomonads consisting of 58 Phl-positive biocontrol strains of worldwide origin and seven Phl-negative representatives of characterized Pseudomonas species were compared using 10 housekeeping genes (i.e. rrs, dsbA, gyrB, rpoD, fdxA, recA, rpoB, fusA, rpsL and rpsG). Multilocus sequence typing differentiated 51 strains among 58 Phl-positive pseudomonads and proved to be as discriminative as enterobacterial repetitive intergenic consensus polymerase chain reaction profiling. As phylogenetic trees inferred from each locus were rather incongruent with one another, we derived the topology from all concatenated loci, which led to the identification of six main groups of Phl-producing Pseudomonas spp. Taxonomically, these groups could correspond to at least six different species. Linkage disequilibrium analysis pointed to a rather clonal structure, even when the analysis was restricted to Phl-producing pseudomonads from a same geographic location or a same phylogenetic group. Intragenic recombination was evidenced for gyrB, rpoD and fdxA, but was shown to be a weaker force than mutation in the origin of intragenetic diversity. This is the first multilocus assessment of the phylogeny and population structure of an ecologically important bacterial group involved in plant disease suppression.
  Environmental Microbiology 09/2007; 9(8):1939-55. · 5.84 Impact Factor
• Article: Characterization and early detection of grapevine (Vitis vinifera) stress responses to esca disease by in situ chlorophyll fluorescence and comparison with drought stress

  Danilo Christen, Susan Schönmann, Mauro Jermini, Reto J. Strasser, Geneviève Défago
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  ABSTRACT: Esca disease, as well as other trunk diseases of grapevine, is an important wood disease that impedes the water transport in plants by clogging the xylem vessels. This type of effect is not detectable for years, due to the long latency time of the disease. In a field experiment, the susceptibility of Vitis vinifera cv. Cabernet sauvignon and Merlot to esca disease was evaluated by visual assessment of foliar symptoms and by necrosis and white rot indexes. C. sauvignon was highly susceptible and Merlot was tolerant to esca. The characteristics of fast chlorophyll a fluorescence transient were investigated in attached leaves by using the so-called JIP-test. The fluorescence transient was analyzed and plants without visible esca foliar symptoms were compared with those showing symptoms. In C. sauvignon, alteration of the photosynthetic apparatus could be detected 2 months before the appearance of foliar symptoms in autumn. To our knowledge, this is the first report of early detection of esca disease using a nondestructive method. For Merlot, only one JIP-test parameter was affected. However, when both cultivars were compared, the relationship of the performance index (defined by the density of reaction centers (RCs) and by the yields φPo and ψo; PIABS = [RC/ABS][φPo/(1 − φPo)][ψo/(1 − ψo)]) versus the calculated rate of the electron transfer [probability that an absorbed photon moves an electron further than View the MathML source; φEo = φPoψo = (TRo/ABS)(ETo/TRo) = ETo/ABS = (1 − F0/FM)(1 − VJ)] permitted us to separate the highest performing cultivar Merlot from the susceptible C. sauvignon. Also, the method used allowed us to detect modification of the photosystem II (PS II) performance in greenhouse-grown Riesling × Sylvaner after a drought stress. Finally, the comparison of the fluorescence transients of esca-affected and drought-stressed grapevines provided information on the differentiated functional-behavior patterns of PS II for the two stress types. These results suggest that esca infection cannot simply be interpreted as a water transport deficit through xylem dysfunction, but that other reaction mechanisms in the plants must be considered. The possibility to use fast chlorophyll a fluorescence monitoring as a wood decay early detection tool is discussed.
  Environmental and Experimental Botany 07/2007; 60(3):504-514. · 2.98 Impact Factor
• Article: Elsinochrome A production by the bindweed biocontrol fungus Stagonospora convolvuli LA39 does not pose a risk to the environment or the consumer of treated crops.

  Désirée Boss, Monika Maurhofer, Elsbeth Schläpfer, Geneviève Défago
  [show abstract] [hide abstract]
  ABSTRACT: Biological control as an alternative to chemical pesticides is of increasing public interest. However, to ensure safe use of biocontrol methods, strategies to assess the possible risks need to be developed. The production of toxic metabolites is an aspect which has so far largely been neglected in the risk assessment and the registration process for biocontrol products. We have evaluated the risks of elsinochrome A (ELA) and leptosphaerodione production by the fungus Stagonospora convolvuli LA39, an effective biocontrol agent used against bindweeds. The toxicity of the two metabolites to bacteria, protozoa, fungi and plants was evaluated in in vitro assays. The most sensitive bacteria and fungi were already affected at 0.01-0.07 microM ELA, whereas plants were far less sensitive. Leptosphaerodione was less toxic than ELA. Subsequently, it was investigated whether ELA is present in the applied biocontrol product or LA39-treated bindweed and crop plants. In plants ELA was never detected and in the biocontrol product the ELA concentration was far too low to have toxic effects even on the most sensitive organisms. We conclude that the production of ELA by biocontrol strain LA39 does not pose a risk to the environment or to the consumer.
  FEMS Microbiology Ecology 02/2007; 59(1):194-205. · 3.41 Impact Factor
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  Available from: goffredo.ch

  Article: Is the ability of biocontrol fluorescent pseudomonads to produce the antifungal metabolite 2,4-diacetylphloroglucinol really synonymous with higher plant protection?

  Fabio Rezzonico, Marcello Zala, Christoph Keel, Brion Duffy, Yvan Moënne-Loccoz, Geneviève Défago
  [show abstract] [hide abstract]
  ABSTRACT: The antifungal compound 2,4-diacetylphloroglucinol (Phl) contributes to biocontrol in pseudomonads, but whether or not Phl(+) biocontrol pseudomonads display higher plant-protecting activity than Phl(-) biocontrol pseudomonads remains to be demonstrated. This issue was addressed by assessing 230 biocontrol fluorescent pseudomonads selected from a collection of 3132 bacterial isolates obtained from 63 soils worldwide. One-third of the biocontrol pseudomonads were Phl(+) and almost all Phl(+) isolates also produced hydrogen cyanide (HCN). The only Phl(+) HCN(-) strain did harbor hcn genes, but with the deletion of a 134 bp hcnC fragment corresponding to an ADP-binding motif. Statistical analysis of biocontrol isolate distributions indicated that Phl production ability was associated with superior disease suppression activity in the Pythium-cucumber and Fusarium-tomato pathosystems, but this was also the case with HCN production ability. However, HCN significance was not as strong, as indicated both by the comparison of Phl(-) HCN(+) and Phl(-) HCN(-) strains and by correlation analyses. This is the first population-level demonstration of the higher plant-protecting activity of Phl(+) biocontrol pseudomonads in comparison with Phl(-) biocontrol pseudomonads.
  New Phytologist 02/2007; 173(4):861-72. · 6.64 Impact Factor
• Article: Thiamine-auxotrophic mutants of Pseudomonas fluorescens CHA0 are defective in cell-cell signaling and biocontrol factor expression.

  Christophe Dubuis, Joëlle Rolli, Matthias Lutz, Geneviève Défago, Dieter Haas
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  ABSTRACT: In the biocontrol strain Pseudomonas fluorescens CHA0, the Gac/Rsm signal transduction pathway positively controls the synthesis of antifungal secondary metabolites and exoenzymes. In this way, the GacS/GacA two-component system determines the expression of three small regulatory RNAs (RsmX, RsmY, and RsmZ) in a process activated by the strain's own signal molecules, which are not related to N-acyl-homoserine lactones. Transposon Tn5 was used to isolate P. fluorescens CHA0 insertion mutants that expressed an rsmZ-gfp fusion at reduced levels. Five of these mutants were gacS negative, and in them the gacS mutation could be complemented for exoproduct and signal synthesis by the gacS wild-type allele. Furthermore, two thiamine-auxotrophic (thiC) mutants that exhibited decreased signal synthesis in the presence of 5 x 10(-8) M thiamine were found. Under these conditions, a thiC mutant grew normally but showed reduced expression of the three small RNAs, the exoprotease AprA, and the antibiotic 2,4-diacetylphloroglucinol. In a gnotobiotic system, a thiC mutant was impaired for biological control of Pythium ultimum on cress. Addition of excess exogenous thiamine restored all deficiencies of the mutant. Thus, thiamine appears to be an important factor in the expression of biological control by P. fluorescens.
  Applied and Environmental Microbiology 05/2006; 72(4):2606-13. · 3.83 Impact Factor
• Article: Genetic diversity and biocontrol potential of fluorescent pseudomonads producing phloroglucinols and hydrogen cyanide from Swiss soils naturally suppressive or conducive to Thielaviopsis basicola-mediated black root rot of tobacco.

  Alban Ramette, Yvan Moënne-Loccoz, Geneviève Défago
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  ABSTRACT: Pseudomonas populations producing the biocontrol compounds 2,4-diacetylphloroglucinol (Phl) and hydrogen cyanide (HCN) were found in the rhizosphere of tobacco both in Swiss soils suppressive to Thielaviopsis basicola and in their conducive counterparts. In this study, a collection of Phl+ HCN+Pseudomonas isolates from two suppressive and two conducive soils were used to assess whether suppressiveness could be linked to soil-specific properties of individual pseudomonads. The isolates were compared based on restriction analysis of the biocontrol genes phlD and hcnBC, enterobacterial repetitive intergenic consensus (ERIC)-PCR profiling and their biocontrol ability. Restriction analyses of phlD and hcnBC yielded very concordant relationships between the strains, and suggested significant population differentiation occurring at the soil level, regardless of soil suppressiveness status. This was corroborated by high strain diversity (ERIC-PCR) within each of the four soils and among isolates harboring the same phlD or hcnBC alleles. No correlation was found between the origin of the isolates and their biocontrol activity in vitro and in planta. Significant differences in T. basicola inhibition were however evidenced between the isolates when they were grouped according to their biocontrol alleles. Moreover, two main Pseudomonas lineages differing by the capacity to produce pyoluteorin were evidenced in the collection. Thus, Phl+ HCN+ pseudomonads from suppressive soils were not markedly different from those from nearby conducive soils. Therefore, as far as biocontrol pseudomonads are concerned, this work yields the hypothesis that the suppressiveness of Swiss soils may rely on the differential effects of environmental factors on the expression of key biocontrol genes in pseudomonads rather than differences in population structure of biocontrol Pseudomonas subcommunities or the biocontrol potential of individual Phl+ HCN+ pseudomonad strains.
  FEMS Microbiology Ecology 04/2006; 55(3):369-81. · 3.41 Impact Factor
• Article: A microsatellite based method for quantification of fungi in decomposing plant material elucidates the role of Fusarium graminearum DON production in the saprophytic competition with Trichoderma atroviride in maize tissue microcosms.

  Andreas Naef, Mauro Senatore, Geneviève Défago
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  ABSTRACT: Common PCR assays for quantification of fungi in living plants cannot be used to study saprophytic colonization of fungi because plant decomposition releases PCR-inhibiting substances and saprophytes degrade the plant DNA which could serve as internal standard. The microsatellite PCR assays presented here overcome these problems by spiking samples prior to DNA extraction with mycelium of a reference strain. PCR with fluorescent primers co-amplifies microsatellite fragments of different length from target and reference strains. These fragments were separated in a capillary sequencer with fluorescence detection. The target/reference ratio of fluorescence signal was used to calculate target biomass in the sample. Such PCR assays were developed for the mycotoxin deoxynivalenol (DON)-producing wheat and maize pathogen Fusarium graminearum and the biocontrol agent Trichoderma atroviride, using new microsatellite markers. In contrast to real-time PCR assays, the novel PCR assays showed reliable fungal biomass quantification in samples with differentially decomposed plant tissue. The PCR assays were used to quantify the two fungi after competitive colonization of autoclaved maize leaf tissue in microcosms. Using a DON-producing F. graminearum wild-type strain and its nontoxigenic mutant we found no evidence for a role of DON production in F. graminearum defense against T. atroviride. The presence of T. atroviride resulted in a 36% lower wild-type DON production per biomass.
  FEMS Microbiology Ecology 03/2006; 55(2):211-20. · 3.41 Impact Factor