Phylogenetic diversity of nitrogen-fixing bacteria in mangrove sediments assessed by PCR-denaturing gradient gel electrophoresis

Culture-independent PCR-denaturing gradient gel electrophoresis (DGGE) was employed to assess the composition of diazotroph species from the sediments of three mangrove ecosystem sites in Sanya, Hainan Island, China. A strategy of removing humic acids prior to DNA extraction was conducted, then total community DNA was extracted using the soil DNA kit successfully for nifH PCR amplification, which simplified the current procedure and resulted in good DGGE profiles. The results revealed a novel nitrogen-fixing bacterial profile and fundamental diazotrophic biodiversity in mangrove sediments, as reflected by the numerous bands present DGGE patterns. Canonical correspondence analysis (CCA) revealed that the sediments organic carbon concentration and available soil potassium accounted for a significant amount of the variability in the nitrogen-fixing bacterial community composition. The predominant DGGE bands were sequenced, yielding 31 different nifH sequences, which were used in phylogenetic reconstructions. Most sequences were from Proteobacteria, e.g. alpha, gamma, beta, delta-subdivisions, and characterized by sequences of members of genera Azotobacter, Desulfuromonas, Sphingomonas, Geobacter, Pseudomonas, Bradyrhizobium and Derxia. These results significantly expand our knowledge of the nitrogen-fixing bacterial diversity of the mangrove environment.