Flagella of a plant-growth-stimulating Pseudomonas fluorescens strain are required for colonization of potato roots

Abstract

The role of motility in the colonization of potato roots by Pseudomonas bacteria was studied. Four TnS-induced flagella-less mutants of the plant-growth-stimulating P. fluorescens WCS374 appeared to be impaired in their ability to colonize growing potato roots.Certain fluorescent Pseudomonas isolates are able to increase plant growth (3,10,16,28) or to protect plants against microbial pathogens (17,25). Our research is focused on strains that can be used to eliminate potato yield reductions as observed in Dutch fields in which potatoes are frequently grown (28). These yield reductions can be abolished by treatment of the seed potatoes with particular Pseudomonas strains (3, 16, 27, 28,). Essential for this beneficial effect of the bacteria on plants is the production of fluorescent siderophores, 22) which supposedly enable the Pseudomonas cells to scavenge most of the Fe3`ions from the Fe3+-poor soil, thereby depriving deleterious microorganisms of this essential element. A second factor which is supposed to be essential for efficient protection of the roots against deleterious microorganisms is the delivery of siderophores along the whole root system of the plant, which requires efficient colonization of the potato roots. Although root colonization is very important in nature, virtually nothing is known about it at the molecular level. We studied the role of motility of Pseudomonas bacteria in the colonization of potato roots. MATERIALS ANID METHODSStrains and growth conditions. The potato root isolate Pseudomonas fluorescens WCS374 (7, 9, 10) is resistant to nalidixic acid (25 ,ug/ml). Strain JM3741 is a TnS-marked derivative of WCS374 which was isolated by J. Marugg. It does not differ significantly from the parental strain in its root-colonizing ability, its siderophore production, its growth rate in the complex King B medium or in minimal medium, and its motility (P. A. H. M. Bakker, unpublished data). Escherichia coli CSH52 harboring the mobilizable plasmid pSUP202 (Apr Cmr Kmr Tcr) and strain S17-1 harboring pSUP2021 (=pSUP202 with Tn5 inserted into the gene for tetracycline resistance) were obtained from R. Simon (29). Pseudomonas strains were grown at 28°C and E. ccli strains were grown at 37°C in King B medium (15) under vigorous aeration. For the root colonization assay TnSlabeled strains were cultivated at 28°C for 48 h on King B medium solidified with 1.6% agar and supplemented with kanamycin (50 ,ug/ml), The TnS-labeled strains were resistant to kanamycin (200 ,ug/ml) and streptomycin (200 ,ug/ml) * Corresponding author.(TnS encodes both kanamycin and streptomycin resistance in Pseudomonas species).Isolation of Tn5-induced nonmotile mutants. TnS-induced mutants were obtained by the method of Simon et al. (29) with slight modifications as described by Marugg et al. (22). Briefly, E. coli S17-1 harboring pSUP2021 was mated with P. fluorescens WCS374 for 3 to 4 h at 28°C. Transconjugants were selected on King B agar plates containing 25 ,ug of kanamycin per ml and 20 ,g of nalidixic acid per ml. Colon...

Analysis of Pseudomonas fluorescens F113 genes implicated in flagellar filament synthesis and their role in competitive root colonization

“…Most of the already described non-motile mutants are severely affected in root colonization, especially at sites most distant from the inoculation site (De Weger et al, 1987). We have also observed the predominance of flagellar variants with enhanced surface motility, in the distal parts of the rhizosphere that are not easily reached by the wild-type strain (SanchezContreras et al, 2002).…”

“…Total bacterial proteins and exported proteins were analysed by Western blotting with an antiflagellin antiserum (De Weger et al, 1987) (Fig. 4).…”

“…The defect was larger at sites more distant from the inoculation site, in the root systems formed after the bacterial inoculation (De Weger et al, 1987;Dekkers et al, 1998b; Chin-a-Woeng et al, 2000). Furthermore motility-impaired mutants of Pseudomonas chlororaphis PCL1391 do not reduce the The GenBank/EMBL/DDBJ accession number for the 6?5 kb region sequence reported in this paper is AF399739.…”

“…Reports to date refer to mutants that are either aflagellate (De Weger et al, 1987) or deficient in chemotaxis (de Weert et al, 2002). The mutants obtained in this work are not affected in chemotaxis but they are affected in motility to different degrees.…”

Fis regulates the competitiveness of Pseudomonas putida on barley roots by inducing biofilm formation

“…On the other hand, the movement of bacteria via flagella appears to be an important trait for the recolonization of plant roots (Dennis et al, 2010;Lugtenberg et al, 2001). For instance, non-motile and flagella-less mutants of P. fluorescens are less competitive than wild-type bacteria in root apex colonization (De Weger et al, 1987), and the long-term incubation of P. fluorescens on the roots of alfalfa selects for enhanced-motility variants of the bacterium (Martínez-Granero et al, 2006). Indeed, although the formation of biofilms may protect plants against pathogens, the colonization of root tips by cells with enhanced biofilm formation ability has been shown to be diminished up to five times compared with wild-type cells.…”