In Rhizobium leguminosarum biovar viciae, the nodABC and nodFEL operons are involved in the production of lipo-oligosaccharide signals which mediate host specificity. The structure of these metabolites and those produced in nod mutants links the nodE and nodL genes to specific chemical features of the signal molecules. A nodE-determined, highly unsaturated fatty acid and a nodL-determined O-acetyl substituent are essential for the ability of the signals to induce nodule meristems on the host plant Vicia sativa.

Root nodules of leguminous plants are symbiotic organs in which Rhizobium bacteria fix nitrogen. Their formation requires the induction of a nodule meristem and the formation of a tubular structure, the infection thread, through which the rhizobia reach the nodule primordium. In the Rhizobium host plants pea and vetch, pre-infection thread structures always preceded the formation of infection threads. These structures consisted of cytoplasmic bridges traversing the central vacuole of outer cortical root cells, aligned in radial rows. In vetch, the site of the infection thread was determined by the plant rather than by the invading rhizobia. Like nodule primordia, pre-infection thread structures could be induced in the absence of rhizobia provided that mitogenic lipo-oligosaccharides produced by Rhizobium leguminosarum biovar viciae were added to the plant. In this case, cells in the two outer cortical cell layers containing cytoplasmic bridges may have formed root hairs. A common morphogenetic pathway may be shared in the formation of root hairs and infection threads.

SummaryRhizobium bacteria produce different surface polysaccharides which are either secreted in the growth medium or contribute to a capsule surrounding the cell. Here, we describe isolation and partial characterization of a novel high molecular weight surface polysaccharide from a strain of Rhizobium leguminosarum that nodulates Pisum sativum (pea) and Vicia sativa (vetch) roots. Carbohydrate analysis showed that the polysaccharide consists for 95% of mannose and glucose, with minor amounts of galactose and rhamnose. Lectin precipitation analysis revealed high binding affinity of pea and vetch lectin for this polysaccharide, in contrast to the other known capsular and extracellular polysaccharides of this strain. Expression of the polysaccharide was independent of the presence of a Sym plasmid or the nod gene inducer naringenin. Incubation of R. leguminosarum with labelled pea lectin showed that this polysaccharide is exclusively localized on one of the poles of the bacterial cell. Vetch roots incubated with rhizobia and labelled pea lectin revealed that this bacterial pole is involved in attachment to the root surface. A mutant strain deficient in the production of this polysaccharide was impaired in attachment and root hair infection under slightly acidic conditions, in contrast to the situation at slightly alkaline conditions. Our data are consistent with the hypothesis that rhizobia can use (at least) two mechanisms for docking at the root surface, with use of a lectin-glycan mechanism under slightly acidic conditions.

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small bacteriocin was isolated from the culture broth of the gram-negative bacterium Rhizobium leguminosarum, which forms symbiotic nitrogen-fixing root nodules on a number of leguminous plants. The structure of the molecule was elucidated by spectroscopic methods and identified as N-(3R-hydroxy-7-cis-tetradecanoyl)-L-homoserine lactone. The absolute configuration of both asymmetric carbon atoms in the molecule was determined by the use of the chiral solvating agents S-(؉)-and R-(؊)-2,2,2-trifluoro-1-(9-anthryl)-ethanol. small bacteriocin is structurally related to the quorum sensing co-transcription factors for genes from other bacteria such as Vibrio fischeri, Pseudomonas aeruginosa, Erwinia carotovora, and Agrobacterium tumefaciens which are involved in animal-microbe or plant-microbe interactions. The mechanism of regulation of such interactions by this kind of co-transcription factors is still unknown in R. leguminosarum.small bacteriocin (small) is produced by strains of all three biovars of Rhizobium leguminosarum and inhibits the growth of R. leguminosarum bv. viciae 248 and several other strains of this species which, like strain 248, contain a self-transmissible plasmid (12,17). Two genes located close to the transfer (tra) genes (17) on the Sym plasmid pRL1JI of strain 248 are responsible for the fact that this strain does not produce small (rps [repression production small] gene) and that it is sensitive to small (sbs [small bacteriocin sensitivity] gene) (12,17). When strain 248 is cured of its Sym plasmid, pRL1JI (as, for example, in strain RBL1390), it is insensitive to small and also produces small (12, 17). To our knowledge, small is produced only by R. leguminosarum strains, and in nonproducing strains of this species, a gene for small production like that in strain 248 is present (12,17). Therefore, the presence of small is considered a characteristic of this species. Since another typical property of this species is symbiotic root nodule formation on certain leguminous plants, it is possible that both properties are related. However, a strain with a Tn5 insertion in the small gene(s) could induce formation of normal root nodules, which shows that the small gene is not required for root nodule formation (16). However, this does not exclude an ecological link between small production and the interaction of Rhizobium spp. with plants, since other genes may complement the lost function or the function is not essential. In order to detect its biological significance, small was extracted from the bacterial culture medium with chloroform (16) and identified chemically. The small molecule appeared to contain an N-acyl homoserine lactone structure, as known from quorum sensing signal molecules, which function as co-transcription factors in bacteria that often interact with higher organisms (reviewed by Fuqua et al. [10]). The implications of this finding are discussed. MATERIALS AND METHODSBacterial strains and growth conditions. The bacterial strains used in this study were the small-sensitive strain...