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Tn5-induced mutants of Rhizobium meliloti that require the amino acids isoleucine and valine for growth on minimal medium were studied. In one mutant, 1028, the defect is associated with an inability to induce nodules on alfalfa. The Tn5 mutation in 1028 is located in a chromosomal 5. Rhizobium meliloti induces nitrogen (N2)-fixing nodules on the roots of alfalfa (Medicago sativa). This symbiotic relationship begins on the root surface, proceeds with the invasion of root hairs, and culminates in the development of an effective N2-fixing nodule. R. meliloti nodulation (nod) genes are located on a large symbiotic plasmid termed pSym (6, 37) and have been cloned and characterized. The common nodulation genes (nodABC) are structurally and functionally conserved among several Rhizobium and Bradyrhizobium species (23,27). Other nodulation genes, such as nodFEGHPQ, are believed to confer specificity for nodulation (12, 21). It has been proposed that the common genes nodABC and the specific genes nodHPQ produce alfalfaspecific signals that trigger a host response (16,26,38). The nodD gene codes for a protein that activates the expression of the other nod genes in the presence of inducing compounds exuded by alfalfa seeds or roots (20,30 In addition, uncharacterized auxotrophic mutants with altered symbiotic properties have been described (14). In this study we have examined the symbiotic properties of two Ilv-mutants of R. meliloti and found that in only one strain the same mutation (ilvC) causes the mutant to be symbiotically defective. We have demonstrated that the mutation in the llv-Nod-mutant maps in the gene coding for the enzyme acetohydroxy acid isomeroreductase (isomero-* Corresponding author. reductase) (E.C. 1.1.1.89) (ilvC). We have determined the primary sequence of the R. meliloti ilvC gene and have shown that the Escherichia coli ilvC gene complements both defective phenotypes in the Ilv-Nod-mutant of R. meliloti. MATERIALS AND METHODSBacterial strains, plasmids, and media. The strains and plasmids used in this study are listed in Table 1. E. coli strains were maintained on LB agar. R. meliloti was grown on either TY medium or minimal medium (28).Amino acids and precursors were purchased from Sigma Chemical Co. unless stated otherwise and were used as netessary in the following concentrations (milligrams per liter): isoleucine, 50; valine, 50; a-ketobutyrate, 125; a-ketoisovalerate, 90; calcium D-pantothenate, 8.4; racemic acetolactate

Rhizobium tropici CIAT899 displays intrinsic tolerance to acidity, and efficiently nodulates Phaseolus vulgaris at low pH. By characterizing a gshB mutant strain, glutathione has been previously demonstrated to be essential for R. tropici tolerance to acid stress. The wild-type gshB gene region has been cloned and its transcription profile has been characterized by using quantitative real-time PCR and transcriptional gene fusions. Activation of the gshB gene under acid-stress conditions was demonstrated. gshB is also induced by UV irradiation. Upstream from gshB a putative s 70 promoter element and an inverted repeat sequence were identified, which are proposed to be involved in expression under neutral and acidic conditions, respectively. Gel retardation assays indicate that transcription in acid conditions may involve protein binding to an upstream regulatory region.