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Title: THE ROLE OF DSBA IN ROOT COLONIZATION BY PSEUDOMONAS FLUORESCENS Q8R1-96. APPLIED AND ENVIRONMENTAL MICROBIOLOGY.

Author
item MAVRODI, O - WASHINGTON STATE UNIV.
item MAVRODI, D - WASHINGTON STATE UNIV.
item Weller, David
item Thomashow, Linda

Submitted to: Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/25/2005
Publication Date: 1/1/2006
Citation: Mavrodi, O.V., Mavrodi, D.M., Weller, D.M., Thomashow, L.S. 2006. The role of dsba in root colonization by pseudomonas fluorescens q8r1-96. Microbiology 152: 863-872.

Interpretive Summary: Root diseases caused by soilborne pathogens are major yield-limiting factors in the production of food, fiber and ornamental crops. Trends in agriculture toward greater sustainability and less dependence on synthetic fungicides have created a need for more ecologically sound methods of disease control. Biological control exploits the natural antagonistic activity of certain root-colonizing bacteria against plant pathogens and is considered the best alternative for the control of certain soilborne root diseases of wheat. This study focused on Pseudomonas fluorescens Q8r1-96, a bacterial strain that colonizes the roots of wheat more aggressively and maintains protective populations longer than do other biocontrol agents. The specific goal was to determine the role of the gene dsbA in the exceptional root-colonizing ability of Q8r1-96. The dsbA gene is required for proper folding of many proteins on the outside of bacterial cells, and such proteins are likely to participate in interactions between Q8r1-96 and its plant host. The dsbA gene in Q861-96 was identified, cloned, characterized, and used to construct a derivative strain with a nonfunctional dsbA gene. This mutant had altered surface properties and was less motile than the parental strain. Unexpectedly, it produced more of the inhibitory compound DAPG than the parental strain; the reason for this is not known. When introduced into soil, the mutant colonized the roots of wheat as well as the parental strain, indicating that the altered surface properties did not greatly affect competitiveness against other soil bacteria. However, the mutant was less competitive than the parental strain when the two were introduced into soil together, suggesting that dsbA is a factor in overall fitness. We conclude from this work that dsbA and, by extension, proteins on the cell surface are unlikely to be major determinants of the exceptional root-colonizing activity of P. fluorescens Q8r1-96.

Technical Abstract: Fluorescent Pseudomonas spp. that produce 2,4-diacetylphloroglucinol 2,4-(DAPG) suppress a wide range of diseases caused by soilborne plant pathogens and have been divided into 17 genotypes (A-Q) by rep-PCR using the BOXA1R primer (BOX-PCR). The genotype of an isolate is predictive of its ability to colonize the rhizosphere of certain crop species. Pseudomonas fluorescens Q8r1-96, a representative of genotype D, is known for its exceptional ability to colonize the roots of wheat and pea as compared to strains of other genotypes. Clones containing the dsbA gene, encoding a periplasmic disulfide bond-forming protein, were identified by Southern hybridization in a Q8r1-96 genomic library, sequenced, and used to construct a gene replacement mutant of Q8r1-96 in order to determine the role of this gene in rhizosphere colonization. The dsbA mutant exhibited decreased motility and fluorescence, and altered colony morphology; however, it produced more 2,4-DAPG and total phloroglucinol-related compounds and inhibited Gaeumannomyces graminis var. tritici in vitro better than did the parental strain. When applied individually into a natural soil, Q8r1-96 and the dsbA mutant did not differ in ability to colonize the rhizosphere of wheat in greenhouse experiments lasting 12 weeks. However, when the two strains were co-inoculated into the soil, the parental strain consistently outcompeted the dsbA mutant. This study demonstrates that dsbA does not contribute to the exceptional rhizosphere competence of Q8r1-96, but reduces its competitiveness only when it must compete with itself in the same niche in the rhizosphere.