GENOMIC REQUIREMENTS OF RHIZOBIUM FOR NODULATION OF WHITE CLOVER HAIRY ROOTS TRANSFORMED WITH THE PEA LECTIN GENE

Soil bacteria from the genera Rhizobium, Bradyrhizobium, or Azorhizobium induce formation of nitrogen-fixing nodules in legume roots in a host plant-specific way. For instance, R. leguminosarum bv. viciae (R. l. bv. viciae) nodulates pea but not white clover roots. Previously, we have shown that introduction of the pea lectin gene (ps1) into white clover hairy roots enabled nodulation of such roots by R. l. bv. viciae (C. L. Diaz, L. S. Melchers, P. J. J. Hooykaas, B. J. J. Lugtenberg, and J. W. Kijne, Nature 338:579-581, 1989). To establish which nodulation (nod) genes of R. l. bv. viciae are required for this heterologous nodulation, we inoculated psl-transformed hairy roots with a set of derivatives of R. l. bv. viciae strain 248, each carrying a transposon insertion in one nod gene as well as with a set of strain 248-derivatives containing a curtailed Sym plasmid pRL1JI carrying nod FDABCIJ and in addition harboring different nod genes cloned in IncP plasmids. The results show that together with the nodD and the nodABCIJ operons, the complete nodFEL operon of R. l. bv. viciae is required for nodulation of white clover hairy roots transformed with the psl gene. The nodulation frequencies and number of nodules observed are comparable to those obtained with strains containing the complete Sym plasmid pRL1JI, regardless of the chromosomal background. Inoculation with the more distantly related R. meliloti and R. etli species did not result in nodule induction. These results indicate that nodulation of psl-transformed white clover hairy roots by R. l. bv. viciae does not represent an aberrant type of nodulation, and involves recognition by white clover of mitogenic R. l. bv. viciae-Nod signals. Interestingly, the homologous biovar R. l. bv. trifolii consistently induced formation of more nodules on psl-transformed white clover hairy roots, which suggests that pea lectin recognizes R. l. bv. viciae as well as its close relative R. l. bv. trifolii.