Diversity of Indigenous Symbiotic Nitrogen Fixing Bacteria from Blackgram [Vigna mungo (L.) Hepper] Cultivated in Rice Fallows

Legume nodulating bacteria (LNB), also known as rhizobia produce root nodules on leguminous plants where in nitrogen fixation takes place. In agriculture, this symbiotic relationship is significant as it provides nitrogenous fertilizers to the leguminous crops. Traditionally rhizobia were exclusively members of the Rhizobiaceae family in the Alphaproteobacteria class of bacteria. Several reports revealed that large number of diazotrophs have the ability to nodulate and fix atmospheric N₂ in legumes in addition to rhizobia which are outside the Alphaproteobacteria class. Relatively less information is available on the genetic diversity of indigenous rhizobia nodulating blackgram cultivated in rice fallows. Hence the present study was carried out to know the diversity of symbiotic native nitrogen fixing bacteria isolated from the root nodules of blackgram grown in rice fallows using partial 16S rRNA sequencing. Nitrogen fixing microsymbionts (Rhizobium strains) from root nodules of blackgram were isolated on yeast extract mannitol agar (YEMA) medium. The pure cultures were maintained at 28±2°C for 48 h. Identification of Rhizobium isolates was done by using various tests and Morphological characterization of isolates was also done by using standard microbiological methods. The PCR amplification and sequencing of 16S rRNA gene of isolates were carried out to identify the isolates. In the present study four strains of Rhizobium designated as VM-2, VM-8, VM-9 and VM-15 were isolated. Morphologically colonies of all strains are round, transparent, entire, convex and smooth. They are Gram -ve and rod shaped. Mucilage production is noticed in VM-2, VM-8 and VM-9. The 16S rRNA gene sequences revealed that the strain VM-2 showed a close relation with Rhizobium, VM-8 and VM-9 with Bradyrhizobium where as VM-15 with Achromobater. It was concluded that the symbiotic nitrogen fixing bacteria isolated from the root nodules of blackgram cultivated in rice fallows exhibited high genetic diversity.