A total of 137 soilborne and plant-associated bacterial strains belonging to different Pseudomonas species were tested for their ability to synthesize N-acyl-homoserine lactones (NAHL). Fifty-four strains synthesized NAHL. Interestingly, NAHL production appears to be more common among plant-associated than among soilborne Pseudomonas spp. Indeed, 40% of the analyzed Pseudomonas syringae strains produced NAHL which were identified most often as the short-chain NAHL, N-hexanoyl-L-homoserine lactone, N-(3-oxo-hexanoyl)-homoserine lactone, and N-(3-oxo-octanoyl)-L-homoserine lactone (no absolute correlation between genomospecies of P. syringae and their ability to produce NAHL could be found). Six strains of fluorescent pseudomonads, belonging to the species P. chlororaphis, P. fluorescens, and P. putida, isolated from the plant rhizosphere produced different types of NAHL. In contrast, none of the strains isolated from soil samples were shown to produce NAHL. The gene encoding the NAHL synthase in P. syringae pv. maculicola was isolated by complementation of an NAHL-deficient Chromobacterium mutant. Sequence analysis revealed the existence of a luxI homologue that we named psmI. This gene is sufficient to confer NAHL synthesis upon its bacterial host and has strong homology to psyI and ahlI, two genes involved in NAHL production in P. syringae pv. tabaci and P. syringae pv. syringae, respectively. We identified another open reading frame that we termed psmR, transcribed convergently in relation to psmI and partly overlapping psmI; this gene encodes a putative LuxR regulatory protein. This gene organization, with luxI and luxR homologues facing each other and overlapping, has been found so far only in the enteric bacteria Erwinia and Pantoea and in the related species P. syringae pv. tabaci.

Fluorescent pseudomonads have evolved an efficient strategy of iron uptake based on the synthesis of the siderophore pyoverdine and its relevant outer membrane receptor. The possible implication of pyoverdine synthesis and uptake on the ecological competence of a model strain (Pseudomonas fluorescens C7R12) in soil habitats was evaluated using a pyoverdine minus mutant (PL1) obtained by random insertion of the transposon Tn5. The Tn5 flanking DNA was amplified by inverse PCR and sequenced. The nucleotide sequence was found to show a high level of identity with pvsB, a pyoverdine synthetase. As expected, the mutant PL1 was significantly more susceptible to iron starvation than the wild-type strain despite its ability to produce another unknown siderophore. As with the wild-type strain, the mutant PL1 was able to incorporate the wild-type pyoverdine and five pyoverdines of foreign origin, but at a significantly lower rate despite the similarity of the outer membrane protein patterns of the two strains. The survival kinetics of the wild-type and of the pyoverdine minus mutant, in bulk and rhizosphere soil, were compared under gnotobiotic and non-gnotobiotic conditions. In gnotobiotic model systems, both strains, when inoculated separately, showed a similar survival in soil and rhizosphere, suggesting that iron was not a limiting factor. In contrast, when inoculated together, the bacterial competition was favorable to the pyoverdine producer C7R12. The efficient fitness of PL1 in the presence of the indigenous microflora, even when coinoculated with C7R12, is assumed to be related to its ability to uptake heterologous pyoverdines. Altogether, these results suggest that pyoverdine-mediated iron uptake is involved in the ecological competence of the strain P. fluorescens C7R12.

The diversity of the membrane-bound nitrate reductase (narG) and nitrous oxide reductase (nosZ) genes in fluorescent pseudomonads isolated from soil and rhizosphere environments was characterized together with that of the 16S rRNA gene by a PCR-restriction fragment length polymorphism assay. Fragments of 1,008 bp and 1,433 bp were amplified via PCR with primers specific for the narG and nosZ genes, respectively. The presence of the narG and nosZ genes in the bacterial strains was confirmed by hybridization of the genomic DNA and the PCR products with the corresponding probes. The ability of the strains to either reduce nitrate or totally dissimilate nitrogen was assessed. Overall, there was a good correspondence between the reductase activities and the presence of the corresponding genes. Distribution in the different ribotypes of strains harboring both the narG and nosZ genes and of strains missing both genes suggests that these two groups of strains had different evolutionary histories. Both dissimilatory genes showed high polymorphism, with similarity indexes (Jaccard) of between 0.04 and 0.8, whereas those of the 16S rRNA gene only varied from 0.77 to 0.99. No correlation between the similarity indexes of 16S rRNA and dissimilatory genes was seen, suggesting that the evolution rates of ribosomal and functional genes differ. Pairwise comparison of similarity indexes of the narG and nosZ genes led to the delineation of two types of strains. Within the first type, the similarity indexes of both genes varied in the same range, suggesting that these two genes have followed a similar evolution. Within the second type of strain, the range of variations was higher for the nosZ than for the narG gene, suggesting that these genes have had a different evolutionary rate.Denitrification is a microbial process in which oxidized nitrogen compounds are used as alternative electron acceptors for energy production when oxygen is limited. Denitrification consists of four reactions by which nitrate is reduced to dinitrogen by the metalloenzymes nitrate reductase, nitrite reductase, nitric oxide reductase, and nitrous oxide reductase. Bacteria capable of denitrification are frequently isolated from soil environments (30). The most common denitrifiers isolated from temperate soils belong to the group of fluorescent Pseudomonas spp. (8).Zones of high denitrifying activity occur predominantly in specific soil microsites (21) and in "activation sites," such as rhizospheric soil (15). Indeed, the proportion of fluorescent pseudomonads able to reduce nitrates appeared to be significantly higher in the rhizosphere than in the bulk soil. Even more, this proportion increases gradually and significantly in the vicinity of root (4). This observation has been made in the rhizosphere of different plant species cultivated in different soils (5). These results indicate that bacteria able to dissimilate nitrogen are selected in the rhizosphere. Furthermore, the nitrate reductase encoded by narG was recently shown to be involved not only in the ...

The lack of consistency of the beneficial effects of inoculated fluorescent pseudomonads has often been related to their bad survival in the rhizosphere. In this review, we describe the strategy followed over the last decade to study traits involved in the rhizosphere competence of these bacteria. The diversity of indigenous populations associated with plant roots was first compared to that of populations associated with uncultivated soils in order to identify traits that discriminate these populations. The involvement of these bacterial traits in the rhizosphere competence was then assessed by comparing the competitiveness of a wild-type strain to that of mutants affected in the corresponding phenotypes. Finally, traits shared by populations adapted to the rhizosphere were identified by comparing both the competitiveness in the rhizosphere and the metabolism of a collection of bacterial strains. The data yielded indicated that rhizosphere-competent pseudomonads show a specific metabolism especially characterized by the efficiency of the pyoverdine-mediated iron uptake and by the ability to reduce nitrogen oxides. population / diversity / model strain / mutant / metabolism Résumé-Identification de caractères impliqués dans la compétence rhizosphérique des Pseudomonas spp. fluorescents : description d'une stratégie basée sur des études de populations et de souche modèle. Le manque de fiabilité des effets bénéfiques déterminés par les Pseudomonas spp. fluorescents inoculés a souvent été attribué à leur mauvaise survie dans la rhizosphère. Au cours de cette synthèse, nous décrivons la stratégie suivie lors des dix dernières années pour étudier les caractères impliqués dans la compétence rhizosphérique de ces bactéries. La diversité des populations indigènes associées aux racines a d'abord été comparée à celle des populations associées à des sols nus. L'implication de ces caractères bactériens dans la compétence rhizosphérique a ensuite été évaluée en comparant la compétitivité d'une souche modèle à celle de mutants affectés dans les phénotypes correspondants. Finalement, des caractères partagés par les populations adaptées à la rhizosphère ont été identifiés en comparant la compétitivité rhizosphérique et le métabolisme d'une collection de souches. Les données recueillies indiquent que les Pseudomonas spp. fluorescents adaptés à la rhizosphère présentent un métabolisme spécifique caractérisé, en particulier, par l'efficacité de leur système d'acquisition du fer basé sur les pyoverdines et par leur aptitude à réduire les oxydes d'azote. population / diversité / souche modèle / mutant / métabolisme