Isolation and Characterization of Bacillus Consortia for Plant Growth Promotion in Rice (Oryza Savita L.)

Rice is one of the most important food crops in the world and staple food for more than 50 % of the global population. Meeting the increasing global demand for food, chemical fertilizers are used on a large scale these days in agricultural practices. Excessive use of chemical fertilizers is leading to diminishing reserve of high quality raw material and increasing energy costs. Hence, use of Bacillus consortia in the form of biofertilizers can be considered as an important strategy. Bacillus represent a high proportion of the soil microbial biomass. Plant rhizospheric soils are a major habitat for Bacillus. The association of rhizospheric Bacillus with plants is found to confer many advantages, such as the nitrogen fixation, phosphate solubilization, potassium mobilization as well as production of antimicrobials, phytohormones, ammonia production, production of 1- Aminocyclopropane-1- carboxylate deaminase (ACC), siderophore production, and plant protection against abiotic and biotic stresses. Therefore, Bacillus inoculants can be used to reduce dependence on chemical fertilizer which would lead towards sustainable agriculture. The enhanced Bacillus dynamics in turn increased plant growth parameter, dry biomass and rice yield as well as soil health due to more availability of nutrients and plant growth promoting substances, thus satisfying all the criteria for sustainability. This book will be of use and interest to consultants, researchers, libraries, entrepreneurs, manufacturers of biofertilizer and for those who wants to venture in to this field. I am also confident that this manual is useful for the extension of Bacillus consortia as a biofertilizer usage to a large number of farmers for improvement of crop yield and other benefits, while minimizing environmental pollution from agrochemical inputs.

Dr. Krunal G. Modi has completed his Bachelor of Science in Biotechnology from Veer Narmad South Gujarat University in 2009. He has completed his Master of Science in Biotechnology from Veer Narmad South Gujarat University in 2011. He hs served as a Teaching Assistant at C.G. Bhakta Institute of Biotechnology from 2011-2013. He then selected as an Assistant Professor (Plant Biotechnology), Navsari Agricultural University, Gujarat in 2013. Presently he is working in the area of plant, microbe, and environment interaction for past 10 years. He has developed Bacillus consortia for plant growth promotion and also decipher the mechanism, how bacteria cleans up environment and degrades fruit waste which could be in long-term used as a method of choice for clean and sustainable agriculture. He has also worked on biological control of phytopathogens using nanoparticles, heavy metal resistant bacteria for bioremediation of contaminated soil. Dr. Modi has published 13 research papers and 11 popular articles in National/International Journals and also published 4 practical manuals.

Dr. Sanjay Jha has completed his Bachelor of Science in Agriculture from GB Pant University in 1997. He has completed his Master of Science in Agril. Biotechnology from Assam Agricultural University in 2001. He has completed his Doctorate in Microbiology from The Maharaja Sayajirao University of Baroda in 2008. Dr. Sanjay Jha has been selected as an Assistant professor (Plant Biotechnology) at Navsari Agricultural University, Gujarat in 2008. He then selected as an Associate Professor (Plant Biotechnology) at Navsari Agricultural University, Gujarat in 2012. Presently he is working in the area of plant-microbe interaction for past 12 years. Dr. Jha has published 50 research papers and 10 popular articles in National/International Journals. Dr. Jha has also guided 6 Ph.D. students and 6 M.Sc. Students.

Dr. Swati Patel has completed her Bachelor of Science in Biotechnology in 2008 from Veer Narmad South Gujarat University. She has completed her Master’s in Biotechnology in 2010 from Veer Narmad South Gujarat University. She has served as a Teaching Assistant at C.G. Bhakta Institute of Biotechnology from 2010-2011. She has achieved her Doctorate in subject Plant Molecular Biology and Biotechnology in 2016 from Navsari Agricultural University, Navsari, Gujarat. Presently she is working on soil conditioning micro-organisms and its interaction for growth promotion and disease resistance for the last 10 years. Dr. Swati Patel has joined as an Assistant Professor in Agricultural Biotechnology at Navsari Agricultural University, Gujarat in 2016. She has published 8 research papers, 13 popular articles in National and International Journals, and 4 practical manuals

1. Introduction........................................................................................
2. Review of Literature ......................................................................... 3

• 2.1 Sustainable agriculture ........................................................ 3
• 2.2 Importance of Rice as a major cereal crop ...................... 4
• 2.3 Rice growth promoting rhizobacteria-the redeemer of plants ........................... 8
• 2.4 Nitrogen fixing bacteria (NFB) in nitrogen nutrition..... 9
• 2.5 Phosphate solubilizing bacteria (PSB) in phosphorus nutrition ............................... 12
• 2.6 Potassium mobilizing bacteria (PSB) in potassium nutrition ............................. 15
• 2.7 Plant growth promoting bacteria in micronutrient nutrition .................... 18
• 2.8 Plant growth regulators .................................................... 19
• 2.9 Siderophore production .................................................... 21
• 2.10 Biological control of fungal rice pathogens ................... 23
• 2.11 PGPB and chemical fertilizers for rice production ....... 27
• 2.12 Bacillus: the protagonist of crop productivity ............... 29
• 2.13 Solutions for enhancement of crop productivity .......... 32
• 2.14 Manipulation of rhizobacteria by recombinant DNA technology................... 34 

3. Materials and Methods................................................................... 37

• 3.1 Isolation, purification, and identification of NFB, PSB, and KMB............................... 37
• 3.2 Quantitative estimation of NFBs ..................................... 39
• 3.3 Secondary screening of PSBs ............................................ 40
• 3.4 Secondary screening of KMBs ......................................... 40
• 3.5 Estimation of organic acid by HPLC .............................. 41
• 3.6 Biochemical characterization of NFB, PSB, & KMB .... 41
• 3.7 Compatibility test ............................................................... 42
• 3.8 Antagonistic activities against rice pathogenic fungi......................... 43
• 3.9 Molecular study of NPK transporters by qRT-PCR analysis ................................ 43
• 3.10 Field preparation ................................................................ 49
• 3.11 Details of the field experiment ......................................... 49
• 3.12 Plant analysis ...................................................................... 50
• 3.13 Soil analysis ......................................................................... 51
• 3.14 Data analysis ....................................................................... 52

4. Results and Discussion ................................................................... 53

• 4.1 Isolation, characterization and identification of NFB, PSB, and KMB.................... 53
• 4.2 Characterization of rhizobacteria ................................... 55
• 4.3 Identification of selected isolates using Biolog system ............................... 55
• 4.4 Molecular characterization .............................................. 58
• 4.5 Secondary screening of NFB ............................................ 59
• 4.6 Phosphate solubilization ................................................... 62
• 4.7 Potassium mobilization ..................................................... 64
• 4.8 Determination of Organic acid ........................................ 66
• 4.9 Siderophore production .................................................... 66
• 4.10 IAA production .................................................................. 68
• 4.11 Compatibility test ............................................................... 69
• 4.12 Antifungal metabolite production .................................. 69
• 4.13 Molecular analysis .............................................................. 70
• 4.14 Field experiment- Interaction with rice GNR-3 (Oryza sativa).................. 81

5. Summary and Conclusion ...........................................................107

References ...............................................................................111