Comparative Evaluation for Shelf Life of The Inoculants Prepared Under Flask Fermentation Conditions and 50 L Fermentor and Their Performance for Nodulation in Gram

A number of climatic, edaphic and biotic factures influence N₂ fixation. Legume- Rhiozobium symbiosis among nitrogen fixing systems has attracted much attention because of specific interaction of plant and bacterial partners. The uniting of a high quality inoculant material composed of highly competitive and environmentally tolerant micro and macro symbionts, with proper agronomic practices, should assure the maximum return for the investment made on BNF technologies. For large-scale production of Rhizobium inoculants for better crop yields, a controlled environment for the growth of microorganisms is very crucial. The success of Rhizobium, inoculation in legumes is mainly dependent on the effectiveness and competitiveness of the strain used. These are the inherent characters of Rhizobium strains and the population of such inoculated efficient strains on seeds at the time of sowing has given competitive advantage to occupy more nodal sites and nodulation. One of the main problems in the quality of Rhizobium inoculants is the survival of microorganisms during storage. For high quality legume inoculant products, the most important factor is presence of live rhizobia capable of nodulation and N₂- fixation within the target host, and little or no contamination. To study the shelf life of rhizobial inoculant, an efficient strain of Rhizobium sp. (G-10) for gram crop was grown under flask fermentation condition for 72 hours and in 50 L fermentor for 36 hours. Inoculants were prepared by shake flask and fermentor grown Rhizobium incorporated in to sterile as well as non-sterile charcoal powder. The samples from the inoculum were taken aseptically after every 20 days to note the viable cell count. The temperature range during the shelf life study of the inoculants was 18–34^oC. In case of rhizobial inoculants impregnated in sterile carrier the cell count at the initial stage was 2.83 x 10⁸cfu/gm, which was decreased to 5.09 x 10⁶ cfu/gm after 100 days for flask fermentation condition. In case of 50 L fermentor the rhizobial count at the initial stage was 5.03 x 10⁸ cfu/gm and decreased to 12.9 x 10⁶ cfu/gm. More decrease in the amount of rhizobia was seen for inoculants in non-sterile carrier prepared by shake flask and 50 L fermentor. The viable cell count at initial stage was 2.01 x 10⁸ cfu/gm and decreased to 0.79 x 10⁶ cfu/gm for shake flask condition. For 50-L fermentor the viable cell count at initial stage was 4.73 x 10⁸ cfu/gm and it decreased to 0.43 x 10⁶ cfu/gm at the 100^th day of inoculant incubation. This decrease may be due to moisture loss and decrease in temperature during storage. In sterile carrier at the end of 100 days, the level of contamination was found to be very low as compared to non-sterile formulations prepared by shake flask as well as 50 L fermentor. A pot experiment was conducted under glass house conditions using flask and fermentor based inoculants for studying their symbiotic parameters (number of nodules, dry weight of nodules and dry biomass of the plant). Maximum number of nodules per plant (43.77 nodules), maximum dry weight of nodules (22.97 mg) and dry weight of plant (8.0 gm) were observed for plants inoculated with fermentor-based bioinoculants. It is concluded that fermentor based bioinoculant prepared in sterile carrier showed best results.