Inga and Lianas Are Key Players in the Tropical Nitrogen Cycle of Brazilian Atlantic Forest: Insights From Linking Rates of Nitrogen Fixation With DNA Barcoding Root Identification

Legumes are abundant and diverse in neotropical forests, and many have the potential to fix atmospheric di-nitrogen, playing a unique role in tropical forest nutrient cycling. Despite their recognized importance, it remains difficult to link rates of nitrogen fixation occurring in root nodules belowground to the responsible host plant. To overcome this challenge, we use DNA barcodes of root nodules, upon which rates of nitrogen fixation have been quantified, to determine which legumes are fixing nitrogen, and how much, during forest succession in the Atlantic Forest of Bahia, Brazil. We developed a three-locus DNA barcode (rbcL, trnH-psbA, and matK) library for the legume tree canopy species. We then excavated root nodules, measured rates of nitrogen fixation, and compared DNA barcode sequences of root nodules to this library and published sequences using a community-curated matK phylogenetic framework. We identified a diversity of legume taxa actively fixing nitrogen. The majority of fixation (> 50%) however, was found to be due to the genus Inga in secondary forest stands, and in mature forests, two genera of lianas (woody vines) (Macropsycanthus & Dioclea) represented 70% of the observed nitrogen fixation. Inga had the greatest range of nitrogenase efficiency (nitrogen fixation per unit of nodule biomass). This study illustrates the key role of Inga in the recovery of the nitrogen cycle in secondary regrowth in this region, the sustained inputs of nitrogen in mature forests by lianas, and the utility and challenges of DNA barcoding for linking rates of nitrogen fixation to the responsible host plant.