Role ofptsP,orfT, andsssRecombinase Genes in Root Colonization byPseudomonas fluorescensQ8r1-96

Abstract

Pseudomonas fluorescens Q8r1-96 produces 2,4-diacetylphloroglucinol (2,4-DAPG), a polyketide antibiotic that suppresses a wide variety of soilborne fungal pathogens, including Gaeumannomyces graminis var. tritici, which causes take-all disease of wheat. Strain Q8r1-96 is representative of the D-genotype of 2,4-DAPG producers, which are exceptional because of their ability to aggressively colonize and maintain large populations on the roots of host plants, including wheat, pea, and sugar beet. In this study, three genes, an sss recombinase gene, ptsP, and orfT, which are important in the interaction of Pseudomonas spp. with various hosts, were investigated to determine their contributions to the unusual colonization properties of strain Q8r1-96. The sss recombinase and ptsP genes influence global processes, including phenotypic plasticity and organic nitrogen utilization, respectively. The orfT gene contributes to the pathogenicity of Pseudomonas aeruginosa in plants and animals and is conserved among saprophytic rhizosphere pseudomonads, but its function is unknown. Clones containing these genes were identified in a Q8r1-96 genomic library, sequenced, and used to construct gene replacement mutants of Q8r1-96. Mutants were characterized to determine their 2,4-DAPG production, motility, fluorescence, colony morphology, exoprotease and hydrogen cyanide (HCN) production, carbon and nitrogen utilization, and ability to colonize the rhizosphere of wheat grown in natural soil. The ptsP mutant was impaired in wheat root colonization, whereas mutants with mutations in the sss recombinase gene and orfT were not. However, all three mutants were less competitive than wild-type P. fluorescens Q8r1-96 in the wheat rhizosphere when they were introduced into the soil by paired inoculation with the parental strain.Interest in biological control continues to increase in response to public concerns about the use of chemical pesticides and recognition of the need for environmentally benign plant disease control strategies (76). Plant growth-promoting rhizobacteria (PGPR) have potential for development as biopesticides, biofertilizers, or phytostimulants, but only a few such products have been marketed, in part because the performance of introduced strains varies among fields and years. Variable root colonization can contribute significantly to this inconsistency because the introduced bacteria must attain threshold population sizes in the rhizosphere in order to be effective. Considerable research during the past 25 years has been directed toward understanding the biotic and abiotic factors that influence root colonization.Our research has focused on colonization of the wheat rhizosphere by Pseudomonas fluorescens Q8r1-96, which produces the polyketide antibiotic 2,4-diacetylphloroglucinol (2,4-DAPG). 2,4-DAPG-producing strains of P. fluorescens suppress root and seedling diseases of a variety of crops and play a key role in the natural biological control of take-all disease of wheat known as take-all decline (4,19,28,49,50,65,75)....

Temperature-Dependent Expression of phzM and Its Regulatory Genes lasI and ptsP in Rhizosphere Isolate Pseudomonas sp. Strain M18

“…Furthermore, PtsP may regulate phzM indirectly by other regulators. As far as we know, the exact regulation mechanism of PtsP has not yet been clarified completely and may involve the utilization of carbon and nitrogen, rather than a promoter-specific regulatory factor (36,53,58). In this report, we provide evidence that the conversion of PCA to PYO by PhzM activity, especially at 37°C, is activated by LasI and inhibited by PtsP, although the precise regulatory mechanism remains to be further investigated in detail.…”

“…3), suggesting that the las system directly activates phzM by recognizing the putative lux box. In previous studies (36,58), inactivation of ptsP resulted in PYO overproduction by P. aeruginosa PAO1, leading to a significant decrease in the expression of qscR and twofold and threefold increases in the expression of lasI and rhlI, respectively, and suggested that PtsP inhibited PYO production by activating qscR and repressing the las and rhl system in P. aeruginosa. Furthermore, PtsP may regulate phzM indirectly by other regulators.…”

Structural and Functional Analysis of the Type III Secretion System fromPseudomonas fluorescensQ8r1-96

“…Mutants were assayed for growth kinetics in King's B and M9 media, swimming motility, and production of exoprotease, siderophore, and 2,4-DAPG as described earlier (41,42). Longterm rhizosphere colonization assays were performed with wheat Triticum aes- tivum L. cv.…”

“…The interrupted inserts from pCR-T3SG2⌬Pst and pCR-T3SG2⌬Sac were subcloned into pEX18Tc. All pEX18Tc-based plasmids were mobilized in Q8r1-96 from E. coli S17-1(-pir), and double crossover events were selected and verified by PCR as described earlier (42).…”

The Multifactorial Basis for Plant Health Promotion by Plant-Associated Bacteria

“…But many other genes are also likely to be important. For example, Mavrodi et al (45) found that three genes related to pathogenicity in some pseudomonads were important for the superior root colonization of a biocontrol strain. To be sure, much work needs to be done in this area, because there are hundreds of genes per genome that are good candidates for mutational studies based on what we already know about biocontrol.…”