In order to understand the community structure of diazotrophs in red soil and effects of organic manure application on the structure, four nifH gene libraries were constructed: the control (CK), low manure (LM), high manure (lIM), and high manure adding lime (ML). Totally 150 nifH gene clones were screened and grouped into 21 clusters by RFLP analysis. Existence of dominant patterns was observed in all libraries, which counted for over 96% of clones in library HM and about 56~72% in other three libraries. The nifH sequences of the dominant patterns in all libraries were most similar to sequences of the cyanobacteria. nifH genes showed high diversity in red soil, dispersing throughout the nifH clades (alpha-, beta-, and gamma-Proteobactena, Firmicutes, cyanobacteria, Verrucomicrobia, and posited group). Bradyrhizobium and Burkholderia were also important diazotrophs in low fertility soil samples. Low manure treatment increased the diversity of nifH genes compared with CK and high manure treatments. Manure and lime treatment led to obvious community succession. Total N to available P ratio, total carbon, and K concentrations were the main factors affecting the diversity of diazotrophs in red soil.