Use of different 15N labelling techniques to quantify the contribution of biological N2 fixation to legumes

The reliable quantification of the contribution of biological N2 fixation (BNF) is an important priority in studies of the N cycle in low-external input farming systems of the tropics where plant-associated BNF if often the largest N input. We have investigated the effect of using more than one control crop, together with different techniques for labelling the soil with 15N, for the application of the isotope dilution technique to quantify the BNF contribution to a forage legume (Centrosema) and French beans (Phaseolus vulgaris). When soil was amended with 15N-labelled organic matter, the 15N enrichment of the soil mineral N decreased slowly with time but when frequent multiple additions of 15N-labelled (NH4)2SO4 were used the 15N enrichment of this N increased with time. Despite this different behaviour of soil 15N enrichment there were no significant differences between 15N treatments in the estimates of the plant N derived from BNF (%Ndfa) of the Centrosema calculated using either of the two non-N2-fixing control crops. This was attributed to the very high %Ndfa of this legume (>84%), a situation where differences in N uptake patterns between control crops are of little importance. In contrast, in the second experiment, the estimates of %Ndfa of P. vulgaris varied from 33 to 62% depending on which 15N addition treatment or control crop was employed. The two estimates derived from the use of non-nodulating bean as control in the different 15N treatments gave a good agreement, which indicated that this crop had a similar soil-N uptake pattern to the nodulated bean crop. The use of several control crops combined with two techniques of 15N addition which produce contrasting temporal changes in the 15N enrichment of the soil N, is a powerful technique for the reliable quantification of BNF contributions to plant nutrition, since it permits the identification of the most suitable control crop from amongst those utilized.