“Delineating bacterial community structure of rhizosphere metagenome from Kansal Forest”

Abstract

Rhizosphere bacteria play crucial role in numerous biochemical and ecological processes ranging from biomass disintegration, mineralization, biological nitrogen fixation and de nitrification. The present investigation was carried out to delineate the microbial diversity of rhizosphere soil using metagenomic approach. Extraction of metagenomic DNA, PCR amplification and cloning of 16S rRNA genes resulted in library consisting of ~2000 clones. Sequencing of randomly selected 400 clones resulted in 352 unique sequences of 16S rRNA. Analysis of recovered sequences using BlastN (NCBI) revealed that ~47% of these sequences had uncultured representatives, uncultured Proteobacteria constituted ~8% of the total sequence diversity, uncultured acidobacteria comprised 5% of the total diversity among recovered clones. Other minor groups represent 2–5% of the total sequence diversity and include Bacillus and Pseudomonas . In addition, 1–2% of the total sequences also include uncultured Commensalibacter sp., Streptomyces sp., uncultured Gemmatimonadales , Halomonas (Gram-negative), uncultured Burkholderia sp., uncultured Pelomonas and other uncultured species. Phylogeny tree revealed that all sequence can be clustered into twelve main clusters indicating their diverse origin. Further clustering of these clones using QIIME resulted in 300 unique OTUs that constitutes 83.57% of the total sequence diversity, 52 clones were clustered as single OTU making a total 14.21%, while one sequence remain unclassified. At the Genus level, Bacillus, Pseudomonas and Staphylococcus demonstrated abundant presence in this soil. At species level Flavisolibacter tropicus of phylum Bacteriodetes showed maximum abundance followed by Bacillus cereus of phylum Firmicutes and Betaproteobacteria bacterium GR16-43 of phylum Proteobacteria.