Arachis species have potential for enhancing cultivated peanut (Arachis hypogaea L.) germplasm as forages and cover crops. This study's objective was to evaluate a range of Arachis species for biological nitrogen fixation capacity. Several Arachis species are tetraploids, and it has been shown that tetraploidy may play an important role in nodule initiation. Species were first tested under natural field conditions and then in the greenhouse using three Bradyrhizobium strains that had been previously shown to be effective on peanut. Nodule number, nodule weight, nitrogenase activity determined by acetylene reduction, and shoot dry weight were measured as indicators of nitrogen fixation capacity. In the field, tetraploid species produced significantly more nodules than the diploids, but total dry matter accumulation was independent of the number of nodules or rate of fixation. In the greenhouse, no significant differences were observed among the bradyrhizobial strains. Arachis hypogaea and A. monticola showed significantly higher measures of nitrogen fixation capacity for all measured traits than the diploid species. However, autotetraploid plants of A. villosa did not have significantly more nodules than diploids of the same accession; the autotetraploids consistently had higher nitrogenase activity. Arachis pusilla never formed a symbiotic relationship with the bradyrhizobial strains used.