Methodologies for estimating nitrogen transfer between legumes and companion species in agro-ecosystems: A review of 15N-enriched techniques

•15N-enrichment methods to estimate N transfer between plant species are reviewed.•Both indirect and the direct 15N techniques are described.•15N data suggest from 0 to >50% of the N in a non-legume can come from a legume.•Transfer of N also occurs from non-legumes to legumes, but is generally lower.•It was concluded that all commonly used methodologies have limitations.

Methodologies based on 15N-enrichment for estimating the transfer of nitrogen (N) between legumes and companion non-legume species in agro-ecosystems are critically reviewed and classified according to whether they (i) are direct or indirect (ii) are dependent or independent of the measurement of yield and (iii) are estimating transfer of legume N, biologically-fixed legume N, or non-legume N. The two most commonly used methods are the indirect 15N isotope dilution method and the direct 15N shoot-labelling method. Published estimates both within and between agro-ecosystems are very variable, ranging from 0 to >50% of the N present in a companion species estimated to have been derived from a companion legume. The transfer of N also occurs from companion species to legumes, but is generally much less than the opposite flux. Factors conducive to N transfer include inter alia species proximity (i.e. intimacy of root contact), legume pruning, defoliation or death and the length of the transfer period. It was concluded that two methodologies (15N2 exposure and the split-root technique) lack field applicability. New insights into the various mechanisms of N transfer in agro-ecosystems may assist in the selection or development of more appropriate methodologies for estimating N transfer by both above- and below-ground pathways.