Background and aims: The selection of legume species and species mixtures influences agroecosystem nitrogen (N) and carbon cycling. We utilized a fertility gradient to investigate the effects of plant species interactions on biological N fixation of an annual and perennial legume in response to shifting soil resource availability. Methods: Legume N fixation of annual field pea ( Pisum sativum) and perennial red clover ( Trifolium pratense) grown in monoculture and mixtures with oats ( Avena sativa) or orchardgrass ( Dactylis glomerata) was estimated using the 15N natural abundance method across 15 farm fields and we measured six soil N pools ranging from labile to more recalcitrant. Results: Evidence of complementary and facilitative species interactions was stronger for the perennial red clover-orchardgrass mixture than for the annual field pea-oat mixture (N Land Equivalency Ratios were 1.6 and 1.2, respectively). We estimated that the transfer of fixed N from red clover to orchardgrass increased aboveground N fixation estimates by 15% from 33 to 38 kg N ha -1. Despite a more than 2-fold range in soil organic matter levels and more than 3-fold range in labile soil N pools across field sites, the N fertility gradient was not a strong predictor of N fixation. While grass N assimilation was positively correlated with soil N pools, we found only weak, inverse correlations between legume N fixation and soil N availability. In grass-legume mixtures, soil N availability indirectly influenced N fixation through plant competition. Conclusions: These results suggest that increasing diversity of cropping systems, particularly through the incorporation of perennial mixtures into rotations, could improve overall agroecosystem N cycling efficiency.
