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Discussion 6 Discussion Public awareness of problems with food safety is constantly increasing, as is the customer demand for food that has been produced by an integrative and sustainable agriculture. While this situation provides a good chance for biological control to prove its commercial applicability, the biggest bottleneck of the system, the high variability in efficiency, remains to be solved. For this reason research on biological control can improve the practical application of biocontrol by investigation of the underling microbial interactions rather than by screening for other control agents. Lack of knowledge on parameters of epiphytical life in the phyllosphere is slowing down the progress in development of biocontrol products. Biological control of bacterial blight of soybean by the antagonist Pss22d is a good model system to gain more information on these parameters, as it provides a) a highly efficient biological control system and b) significant amount of background information on the pathosystem involved. Thus the active principles of biological control could be directly interconnected to the complex plant-microbe interaction. In the frame of this work it was attempted to elucidate the influence of siderophore production on the above mentioned biocontrol system. The results of this study clearly opposed the initial hypothesis, that competition for iron between pathogen and control organism is an active principle of this biological control. Despite this, we could identify a siderophore production system that had not been described in plant-associated P. syringae before and address the importance of this siderophore for epiphytical fitness of the antagonist could be addressed. The production of this second, achromobactin-like siderophore is widely distributed among the pathovars of P. syringae. 6.1 Identification of an achromobactin-like siderophore in P. syringae Iron uptake is essential for most bacteria, thus it is not surprising that the increasing number of genome sequences reveal more and more information on 75
Discussion bacteria with redundandent siderophore systems. For example, Pseudomonas aeroginosa produces pyoverdin and pyochelin, some strains of P. fluorescense produce pyoverdin and pseudomonin, and D. chrysanthemi produces chrysobactin and achromobactin (Cox & Adams, 1985; Franza & Expert, 1991; Franza et al., 2005; Mercado-Blanco et al., 2001; Persmark et al., 1989). While the production of pyoverdin by different pathovars of P. syringae had been reported previously (Bultreys et al., 2001; Jülich et al., 2001), we are the first to proof experimentally the production of an addtional siderophore aside of pyoverdin. This study demonstrated that the biosynthesis gene cluster of this siderophore showed high similarity towards the achromobactin siderophore gene cluster of D. chrysanthemi (Fig.13). IEF analysis of achromobactin and the second siderophore of Pss22d shows a CAS-active band at the same migration (data not shown). TLC separation of achromobactin and the second siderophore of Pss22d on silica plates using a mobile phase of diethylether, n-butanole, acetic acid, water (1:1:1:1; v/v) exhibited a similar CAS-active band (Fig.19A). More importantely, the siderophore of Pss22d∆Pvd could enhance growth of a siderophore-deficient D. chrysanthemi mutant under iron limiting conditions, while the growth deficiency of a mutant defective in the achromobactin receptor was not restored by feeding with Pss22d∆Pvd supernatant (D. Expert, personal communication). All these results imply a close structural similarity between the novel Pss22d siderophore with achromobactin from D. chrysanthemi. . The genetic organization of the biosynthesis genes differs between P. syringae and D. chrysanthemi. The gene acsF, encoding for a putative aminotransferase, is located upstream of the achromobactin receptor gene in D. chrysanthemi, while it follows downstream this receptor gene in P. syringae. This divergent organization could be the result of a genomic reorganization, however it could also indicate a different origin and therefore a different substrate specificity of the two respective aminotransferases. In support of this, the observed color change of the Pss22d∆Pvd siderophore upon iron addition 76
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Siderophore production of Pseudomon
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Acknowledgements This project was s
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Abbreviations Ach achromobactin-lik
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Contents 4.1.2 Pseudomonas syringae
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Contents 5.3 Influence of sideropho
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Introduction 2 Introduction Like hu
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Introduction In contrast to the rhi
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Introduction 2.1.2 Host, pathogen a
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Introduction The relationship betwe
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Introduction developement of resist
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Introduction Fe 2+ + H 2 O 2 → Fe
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Introduction 1997a; May et al., 199
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Introduction Pss22d demonstrated a
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Introduction 2.4 Aim of this study
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Material and Methods Tab. 4. Antibi
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Material and Methods Iron-free 5b m
Page 33 and 34: Material and Methods Solution3, car
Page 35 and 36: Material and Methods 3.6 Plasmids T
Page 37 and 38: Material and Methods PCR-Screening
Page 39 and 40: Material and Methods on ice for 10
Page 41 and 42: Material and Methods stained in eth
Page 43 and 44: Material and Methods 4.2.5.3 Conver
Page 45 and 46: Material and Methods 4.2.5.9 Ligati
Page 47 and 48: Material and Methods strains were s
Page 49 and 50: Material and Methods 9.5). For sign
Page 51 and 52: Material and Methods gentle shaking
Page 53 and 54: Material and Methods columns were w
Page 55 and 56: Results 5 Results 5.1 Analysis of s
Page 57 and 58: Results The 13-kb open reading fram
Page 59 and 60: Results If integration is due to a
Page 61 and 62: Results Yersiniabactin is a siderop
Page 63 and 64: Results Next, a large number of tra
Page 65 and 66: Results Nevertheless the site of tr
Page 67 and 68: Results Achr2_KpnI_rev binding site
Page 69 and 70: Results Interestingly the mutant Ps
Page 71 and 72: Results XAD4-purification yielded i
Page 73 and 74: Results After changing the mobile p
Page 75 and 76: Results P P A Fig. 21. Isoelectic f
Page 77 and 78: Results reduced in comparison to th
Page 79 and 80: Results A-E represents single inocu
Page 81 and 82: Results Tab. 10. Development of pop
Page 83: Results Tab. 11. Distribution of th
Page 87 and 88: Discussion 6.2 Possible regulation
Page 89 and 90: Discussion stationary phase transit
Page 91 and 92: Discussion and bacterium (Loper et
Page 93 and 94: Literature Bultreys, A., Gheysen, I
Page 95 and 96: Literature Galperin, M. Y. (2006).
Page 97 and 98: Literature Mazzola, M. (2002). Mech
Page 99 and 100: Literature Schubert, S., Rakin, A.,
Page 101: Literature Zou, J., Rodriguez-Zas,
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