Improved methodology for tracing a pulse of 13C-labelled tree photosynthate carbon to ectomycorrhizal roots, other soil biota and soil processes in the field

Isotopic pulse-labelling of photosynthate allows tracing of carbon (C) from tree canopies to below-ground biota and calculations of its turnover in roots and recipient soil microorganisms. A high concentration of label is desirable but is difficult to achieve in field studies of intact ecosystem patches with trees. Moreover, root systems of trees overlap considerably in most forests, which requires a large labelled area to minimize the impact of C allocated below-ground by un-labelled trees. We describe a method which combines a high level of labelling at ambient concentrations of CO2, [CO2], with undisturbed root systems and a model to account for root C and root-derived C from un-labelled trees. We raised 5-m-tall chambers, each covering 50 m2 of ground (volume 250 m3) in a young boreal Pinus sylvestris L. forest with up to 5 m tall trees. Rather than a conventional single release of 13CO2, we used five consecutive releases, each followed by a draw-down period, thus avoiding high [CO2]. Hence, we elevated successively the 13CO2 from 1.1 to 23 atom% after the first release to 61 atom% after the fifth, while maintaining [CO2] below 500 p.p.m. during 4-4.5 h of labelling. The average abundance of 13CO2 was as high as 42 atom%. We used the central 10 m2 of the 50 m2 area for sampling of roots and other soil biota. We modelled the dilution of labelled C across the plots by un-labelled C from roots of trees outside the area. In the central 10 m2 area, ~85% of roots and root-associated biota received C from labelled trees. In summary, we elevated the labelling of roots and associated soil biota four-fold compared with previous studies and described the commonly overlooked impact of roots from un-labelled trees outside the labelled area.