Long-term effects of root removal and liming on non-symbiotic N2 fixation in boreal forests

Biological N₂ fixation is a major nitrogen (N) source in boreal forests, but the factors controlling N₂ fixation in boreal forests are not well understood. Most plants in boreal forests rely on mycorrhizal fungi to take up N, boreal forest soils are typically acidic, and the high abundance of mycorrhizal fungi and low soil pH could restrict the abundance of diazotrophs and N₂ fixation. To investigate the effects of mycorrhizal fungi and soil pH on N₂ fixation two experiments were conducted over several years in a mature Pinus sylvestris forest in the boreal zone of Sweden; a pine and shrub root exclusion experiment and a liming experiment. We measured non-symbiotic N₂ fixation, soil carbon (C) and nutrient availability, and the quality of soil organic matter over one growing season, eight and 40 years after the root trenching and liming experiments started, respectively. Both experiments showed that N₂ fixation was still very low in June (during a 48 h incubation at 15°C), indicating that diazotrophs are dormant during large parts of the years and only regain their activity slowly. Further, we found that exclusion of pine roots and associated ectomycorrhizal fungi significantly increased the rate of non-symbiotic N₂ fixation in the late growing season, while exclusion of shrubs and associated ericoid mycorrhizal fungi showed no significant effect. Exclusion of pine roots, shrubs and the associated mycorrhiza strongly increased soil NH₄⁺-N concentrations and the aromaticity of the water-extractable organic matter but did not significantly affect non-symbiotic N₂ fixation. The reason for this might be that the low soil C quality and the high soil N availability offset the effects of the reduced abundance of mycorrhizal fungi on N₂ fixation. In contrast to our expectation, liming did not significantly increase the rate of N₂ fixation, suggesting that soil pH was not the key factor limiting N₂ fixation. Overall, this study suggests that the diazotrophs are in a dormant state during most part of the year. Further, the results indicate that tree roots and ectomycorrhizal fungi rather than shrubs and ericoid mycorrhizal fungi or acidic conditions restrict non-symbiotic N₂ fixation in boreal forests.