To understand the diversities of diazotrophs and denitrifiers in red paddy soil under long-term fertilization conditions, nifH, nirK and nosZ libraries were constructed based on the PCR-RFLP method. nirK gene diversity proved to be lower than that of nosZ and nifH, and nirK and nosZ genes were more sensitive to different fertilization treatments than those with the nifH gene. Diverse microbes including the α-, β-, γ- and δ- subclasses of the Proteobacteria dominated the three libraries. Long-term addition of urea with straw-mulching and azophoska increased the abundance of non-symbiotic diazotrophs, which indicated that non-symbiotic diazotrophs were responsible for the majority of the nitrogen-fixing ability in paddy soil. In addition, a potential link between nifH and nosZ was found due to the existence of nitrogen fixers, such as Bradyrhizobium and Ralstonia in the nosZ library. The main chemical factors affecting the three genes were identified, pH was the most important factor of nifH community, and nirK genes were more affected by pH and organic matter, available potassium and carbon to nitrogen ratio significantly influenced the community structure of the nosZ genes.
Responses of N2O reductase gene (nosZ)-denitrifier communities to long-term fertilization follow a depth pattern in calcareous purplish paddy soil