During the symbiosis between the bacterium Rhizobium meliloti and plants such as alfalfa, the bacteria elicit the formation of nodules on the roots of host plants. The bacteria infect the nodule, enter the cytoplasm of plant cells and differentiate into a distinct cell type called a bacteroid, which is capable of fixing atmospheric nitrogen. To discover bacterial genes involved in the infection and differentiation stages of symbiosis, we obtained genes expressed at the appropriate time and place in the nodule by identifying promoters that are able to direct expression of the bacA gene, which is required for bacteroid differentiation. We identified 230 fusions that are expressed predominantly in the nodule. Analysis of 23 sequences indicated that only three encode proteins known to be involved in the Rhizobium-legume symbiosis, six encode proteins with homology to proteins not previously associated with symbiosis, and 14 have no significant similarity to proteins of known function. Disruption of a locus that encodes a protein with homology to a cell adhesion molecule led to a defect in the formation of nitrogen-fixing nodules, resulting in an increased number of nitrogen-starved plants. Our isolation of a large number of nodule-expressed genes will help to open the intermediate stages of nodulation to molecular analysis.