Isolation, Biochemical Characterization and Production of Biofertilizer from Bacillus megaterium

The isolation of phosphate solubilizing bacterial strains exhibiting high ability to solubilize soil phosphorus is a matter of great interest with high applicability. The use of phosphate solubilizing bacteria as inoculants simultaneously increases phosphate uptake by the plant and increase crop yield. Strains from the genera Pseudomonas, Bacillus and Rhizobium species are among the most powerful phosphate solubilizers. In this present study different cultivated soil samples were investigated for the isolation of phosphate solubilizing bacteria by Pikovskayas agar media. We were found 2 bacterial strains SS1 and SS2 as a phosphate solubilizing bacteria. Both two bacteria are characterized by morphological and biochemical tests. The strain SS2 was confirmed as a Bacillus megaterium. Then Bacillus megaterium is used for the production of longer sustainable phosphate solbulizing biofertilizer. After an interval of 180 days it has 5x10 CFU count. Key-wordsPhosphate solubilization; Soil bacteria; Plant-growth-promoting bacteria; Rhizobacteria; Phosphates; Biofertilizer -------------------------------------------------IJLSSR----------------------------------------------INTRODUCTION Bacillus megaterium was discovered and described [1] in 1884. Bacillus megaterium was used by Lwoff and Guttman in the studies that discovered lysogeny. Bacillus megaterium is one of the first bacteria's genome that has been fully coded. Bacillus megaterium is a biological fertilizer based on a selected strain of naturally-occurring beneficial eubacteria Bacillus megaterium. Bacillus megaterium is a gram positive, rod shaped, endospore forming bacteria. It is used as an effective soil inoculant. Bacillus megaterium have ability to solubilize phosphorus, which is good for plant. Phosphorus plays a significant role in several physiological and biochemical plant activities like photosynthesis, transformation of sugar to starch, transporting of the genetic traits . Access this article online Quick Response Code: Website: www.ijlssr.com DOI: 10.21276/ijlssr.2016.2.6.16 To achieve a high-yield in agriculture, it is necessary to apply phosphorus fertilizers that deliver the nutrients to plants . However, the production of phosphorus fertilizer is a costly process that requires use of nonrenewable phosphate resource (phosphorite), mineral acids (sulfuric acid) and generates many environmental hazardous byproducts . Phosphorus solubilizing microorganisms are ubiquitous in soils and could play an important role in supplying phosphorus to the plants . The principal mechanism for mineral phosphate solubilization is the production of organic acids, and acid phosphatases play a major role in the mineralization of organic phosphorous in soil by Ahmad Ali Khan . Microorganisms also solubilize sparingly soluble phosphates by decreasing the pH of the surrounding environment or acting on the calcium, iron, aluminum, and magnesium salts. In rice plantations, for example, a large amount of organic acids is generated, this increasing phosphorus availability to wheat . Bacillus megaterium var. phosphaticum was used to create a bio-preparation called Phosphobucterin with the purpose of enhancing mineral phosphorus solubilization . If phosphorus is present in the complex structures of the soil and, at the same time, readily decomposable carbon Research Article (Open access) Int. J. Life. Sci. Scienti. Res., VOL 2, ISSUE 6 http://ijlssr.com Copyright © 2015-2016 International Journal of Life-Sciences Scientific Research Page 750 sources, such as manure, are incorporated in the soil, phosphorus solubilization can be increased due to biological activity stimulation. This organic carbon increase may aid to complexing the soila luminum in acids, thus reducing the aluminum phosphate. Taking into account that the isolation of bacterial strains exhibiting high potential of soil phosphorus solubilization has been little studied in Romania, we believe it would be useful to approach this subject of great interest and practical applicability. Approximately 90% to 95% phosphorus formed in soil is present in insoluble forms , which are not utilized by plants, so phosphorus solubilizing bacteria play an important role to solubilize phosphorus, P. vazquez . MATERIALS AND METHODS This project is performed in Microbiology section of Chaperon Biotech Pvt. Ltd, Kanpur, India for duration of 8 Months. Isolation of Bacillus megaterium bacteria from soil sample was done on Pikovskayas agar media by dilution plate technique. Detection and estimation of the phosphate solublization ability of microorganisms have been possible using plate screening method. Phosphate solubilizers produce clearing zones around the microbial colonies in media. Insoluble mineral phosphates such as tricalcium phosphate or hydroxyapatite are contained in the media. The phosphate solubilizing bacteria is isolated for further identification by morphological & biochemical analysis. The morphological tests performed are Gram Staining & Endospore staining. The biochemical analyses performed were Catalase test, Starch hydrolysis test, Citrate hydrolysis test, Methyl red test, Vogous proskar test, Casein hydrolysis test, Indole test, Gelatine hydrolysis test, Urease hydrolysis test, Mannitol fermentation test, Carbohydrate fermentation test, and Salinity test etc. is shown in Table 1. The biofertilizer is prepared by immobilizing the bacterial cell and kept for 36 hours at 37C on rotatory shaker. After immobilization the bacterial culture is added to carrier i.e. charcoal and preserved for further use. Cultivation of phosphate solubilizing Bacillus