Biological nitrogen fixation in barren soils of a high-vanadium region: Roles of carbon and vanadium

Free-living nitrogen fixation (FLNF) is a vital source of nitrogen for the initiation and development of ecosystems on bare lands. This process is catalyzed by the enzyme nitrogenase and is energy intensive. Of the three nitrogenase isoforms, molybdenum nitrogenase (Mo-Nase) is more efficient than its vanadium (V) and iron (Fe) counterparts at room temperature. However, the acquisition of Mo, one of the scarcest biometals in soils, represents a major resource investment for soil diazotrophs. In a vegetation restoration chronosequence developed on barren (low carbon and nitrogen) soils of a high-V region, we tested the following two hypotheses. First, the FLNF rate would be limited by the supply of energy. Second, high V availability would cut the cost of V acquisition, and it would be beneficial for diazotrophs to use V-Nase as a complement of Mo-Nase. For the soils collected along the chronosequence, the addition of a carbon cocktail remarkably stimulated the FLNF rate by relieving the energy constraint and by enriching the diazotrophic genus Paenibacillus. Despite the high V content in the soils, we failed to detect the vnf genes of V-Nase by metagenomic sequencing. Meanwhile, we observed a limited contribution of V-Nase activity to FLNF by checking the R ratio (ratio of FLNF rates measured by acetylene reduction and 15N2 incorporation) and the production of ethane during acetylene reduction. The reason might be that the Mo availability and temperature in our study area did not reach the thresholds for the onset of V-Nase activity. Overall, our results highlight the importance of carbon supply for FLNF in barren soils, indicating that plants may effectively regulate rhizosphere diazotrophs by secreting root exudates. This may be an important mechanism by which nonnitrogen-fixing plants survive on nitrogen-poor soils at the beginning of primary succession.