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Biocontrol by Phenazine-1-carboxamide-Producing Pseudomonas chlororaphis PCL1391 of Tomato Root Rot Caused by Fusarium oxysporum f. sp. radicis-lycopersici

¹Leiden University, Institute of Molecular Plant Sciences, Clusius Laboratory, Wassenaarseweg 64, 2333 AL Leiden, The Netherlands; ²Utrecht University, Department of Mass Spectrometry, F.A.F.C. Wentgebouw, Sorbonnelaan 16, 3584 CA Utrecht, The Netherlands; ³Núcleo de Pesquisas de Produtos Naturais, Universidade Federal do Rio de Janeiro, Centro de Ciências da Saude, Bloco H, 21941-590-Ilha do Fundão, Rio de Janeiro-RJ, Brazil; ⁴Novartis Seeds BV, Westeinde 62, Postbus 26, 1600 AA Enkhuizen, The Netherlands; ⁵Laboratoire de Biologie Microbienne, Université de Lausanne, CH-1015 Lausanne, Switzerland; ⁶Utrecht University, Department of Plant Ecology and Evolutionary Biology, Section of Plant Pathology, P.O. Box 800.84, 3508 TB Utrecht, The Netherlands; ⁷TüV Energie und Umwelt GmbH, Niederlassung Freiburg, Robert-Bunsen Strasse 1, 79108 Freiburg i. Br., Germany; and ⁸MSU-DOE Plant Research Laboratory, Michigan State University, East Lansing 48824, U.S.A.

Seventy bacterial isolates from the rhizosphere of tomato were screened for antagonistic activity against the tomato foot and root rot-causing fungal pathogen Fusarium oxysporum f. sp. radicis-lycopersici. One isolate, strain PCL1391, appeared to be an efficient colonizer of tomato roots and an excellent biocontrol strain in an F. oxysporum/tomato test system. Strain PCL1391 was identified as Pseudomonas chlororaphis and further characterization showed that it produces a broad spectrum of antifungal factors (AFFs), including a hydrophobic compound, hydrogen cyanide, chitinase(s), and protease(s). Through mass spectrometry and nuclear magnetic resonance, the hydrophobic compound was identified as phenazine-1-carboxamide (PCN). We have studied the production and action of this AFF both in vitro and in vivo. Using a PCL1391 transposon mutant, with a lux reporter gene inserted in the phenazine biosynthetic operon (phz), we showed that this phenazine biosynthetic mutant was substantially decreased in both in vitro antifungal activity and biocontrol activity. Moreover, with the same mutant it was shown that the phz biosynthetic operon is expressed in the tomato rhizosphere. Comparison of the biocontrol activity of the PCN-producing strain PCL1391 with those of phenazine-1-carboxylic acid (PCA)-producing strains P. fluorescens 2-79 and P. aureofaciens 30-84 showed that the PCN-producing strain is able to suppress disease in the tomato/F. oxysporum system, whereas the PCA-producing strains are not. Comparison of in vitro antifungal activity of PCN and PCA showed that the antifungal activity of PCN was at least 10 times higher at neutral pH, suggesting that this may contribute to the superior biocontrol performance of strain PCL1391 in the tomato/F. oxysporum system.

Additional keywords: microbiological control.