Long-term application of lime or pig manure rather than plant residues suppressed diazotroph abundance and diversity and altered community

Biological fixation of atmospheric dinitrogen (N2) is an important process that replenishes biologically available nitrogen (N) in soil and helps minimize the use of inorganic N fertilizer in agricultural ecosystems. Diazotrophs are key fixers of atmospheric N2 in a range of soil types, however, there is uncertainty about how they respond to long-term fertilization. Here, using the nifH gene as a molecular marker, we investigated the long-term effects of inorganic and organic fertilization on diazotroph abundance and community structure in an acidic Ultisol. The field experiment ran for 27 years and comprised seven treatments: no fertilization (control); inorganic NPK fertilizer (N); inorganic NPK fertilizer + lime (CaCO3) (NL); inorganic NPK fertilizer + peanut straw (NPS); inorganic NPK fertilizer + rice straw (NRS); inorganic NPK fertilizer + radish (NR); and inorganic NPK fertilizer + pig manure (NPM). Long-term application of fertilizer reduced the abundance and diversity of nifH gene compared with the control (P < 0.05), while lime and pig manure increased the inhibitory effects (P < 0.05). The abundance and diversity of nifH genes were negatively correlated with soil pH, indicating that increasing soil pH potentially affect N fixation ability in acidic Ultisols. Community structure of diazotrophs in the NPS, NRS, and NR treatments were similar and shared most operational taxonomic units (OTUs) with the N treatment, with only a minor difference to that of the control. Thus, there was no effect of plant residue types on diazotroph community structure. In contrast, the application of inorganic fertilizer + liming or pig manure altered the diazotroph community structure with shifts in the dominant genus, from Bradyrhizobium in the control to Azohydromonas in NL and Azospirillum in NPM. Soil pH was the key factor correlated with change in diazotroph community structure. Overall, our results suggested that regular use of lime or pig manure rather than different types of plant residue reduced the abundance and diversity and altered community structure of diazotrophs, that may potentially affect N fixation ability in acidic Ultisols.