A 5.4 kb BamHl fragment of R. leguminosarum bv. trifolii TA1 was found to carry genes involved in exopolysaccharide synthesis (caro genes). This fragment was strongly hybridized to the total DNA from R. I. bv. viciae and bv. phaseoli digested with EcoRI. No homology was found with total DNA of R. meliloti and Rhizobium sp. NGR 234. The exo genes from R. I. bv. trifoliiTAl conjugally introduced into R. I. bv. viciae 1302 considerably affected the symbiosis: the nodules induced on vetch were abortive and did not fix nitrogen. On the other hand, Phaseolus beans infected with R. I. bv. phaseoli harbouring R. I. bv. trifolii exo genes formed the nitrogen-fixing nodules. It can be concluded that additional copies of exo genes introduced into wild type Rhizobium leguminosarum strains can disturb the synthesis of acidic exo- polysaccharides and affect symbiosis of the plants forming indeterminate nodules, but do not affect symbiosis of the plants forming the determinate nodules.
Rhizobium strains isolated from nodules of the different legumes including wild-growing plants were examined for their siderophore activity. Fifteen of the 84 screened rhizobial strains were able to grow under conditions of limited iron supply. Nine of them gave orange halos in the assay with Chrom azurol S. Among these strains were Rhizobium sp. (Ononis) and Rhizobium (Genista), producing hydroxamates and phenolates. These compounds could promote the growth of siderophore-negative bacteria on iron-deficient media. The results imply that the hydroxamates from G1 and Ol strains may belong to the monohydroxamate class of siderophores.
Rhizobium lexuminosarum bv. trifolii produces an acidic exopolysac- charide (EPS) which plays an important role in the development of nitrogen-fixing nodules. Tn5 mutant of R. trifolii <>3 defective in EPS production (Exo") forms ineffective (Fix-) nodules on red clover. This Exo mutation is complemented by the pARF1368 and pARF25c<>Miiid.s isolated fnuii bank of Rhizobium trifolii TA1, but the complementation is not correlated with restoration of Fix* phenotypt-. Furthermore, these cosmids introduced to wild-type of R. trifolii 24 repress its ability to form nitrogen- -fixing nodules. These results might suggest that bacteria with cosmids carrying the exo region form FPS of altered structure. It has been shown by 'll-n.m.r. that exopolysaccharides produced by R. trifolii 93pARF- 1368 and 93pARF25 contain less non-carbohydrate residues (acetyl, pyruvyl and 3-hydroxybutanoyl) than the wild type KPS. These data suggest that the biological activity of the exopolysaccharide of R. trifolii depends on the contents of the non-carbohydrate substitutions.