A deletion mutant of Rhizobium leguminosarum biovar viciae lacking the host-specific nodulation (nod) gene region (nodFEL nodMNT and nodO) but retaining the other nod genes (nodD nodABCIJ) was unable to nodulate peas or Vicia hirsuta, although it did induce root hair deformation. The mutant appeared to be blocked in its ability to induce infection threads and could be rescued for nodulation of V. hirsuta in mixed inoculation experiments with an exopolysaccharide deficient mutant (which is also Nod-). The nodulation deficiency of the deletion mutant strain could be partially restored by plasmids carrying the nodFE, nodFEL or nodFELMNT genes but not by nodLMN. Surprisingly, the mutant strain could also be complemented with a plasmid that did not carry any of the nodFELMNT genes but which did carry the nodO gene on a 30 kb cloned region of DNA. Using appropriate mutations it was established that nodO is essential for nodulation in the absence of nodFE. Thus, either of two independent nod gene regions can complement the deletion mutant for nodulation of V. hirsuta. Similar observations were made for pea nodulation except that nodL was required in addition to nodO for nodulation in the absence of the nodFE genes. These observations show that nodulation can occur via either of two pathways encoded by non-homologous genes.