Antiphytopathogenic and Plant Growth Promoting Attributes of Bacillus Strains Isolated from Rhizospheric Soil of Chickpea

Authors
S. Patil
C. T. Shivannavar
M. C. Bheemaraddi
S. M. Gaddad
Department of Microbiology, Gulbarga University, Gulbarga-585106, Karnataka, India.
Abstract
In the present study, we aimed to screen indigenous rhizospheric Bacillus strains, capable of producing antiphytopathogenic and plant growth promoting traits. Isolate CTS-B19 and CTS-G24 exhibited quite noticeable antagonistic activity initially against Fusarium oxysporum f. sp. ciceri and Rhizoctonia bataticola, and, later, against a panel of phytopathogens. Partial 16S rRNA sequence analysis showed that the isolate CTS-B19 and CTS-G24 exhibited 99% homology with Paenibacillus polymyxa and Bacillus subtilis and the sequences were deposited in GenBank with accession numbers KF322038 and KF322037, respectively. In vitro detection for fungal wall degrading enzymes revealed that both isolates produced chitinases, β-1,3-glucanases, proteases and cellulases. While siderophores and catalase activities were observed only in Bacillus subtilis (CTS-G24), both strains exhibited a positive result for in vitro ammonia production. Besides, the strain CTS-B19 could also solubilize phosphate. Lytic enzymes and siderophore produced by Bacillus subtilis can be considered as potential antiphytopathogenic traits involved in the inhibition of fungal growth. Both strains exhibited either no or negligible antagonistic activity against other plant growth promoting bacteria. Additional to antagonism, plant growth promoting traits exhibited by these natural biocontrol agents may suppress plant diseases and might be applied in agriculture as an alternative to chemical pesticides and fertilizers.

Keywords

1. Adams, D. J. 2004. Fungal Cell Wall Chitinase and Glucanases: Mini-Review. Microbiol., 150: 2029-2035.
2. Bashan, Y. 1998. Inoculants of Plant Growth-promoting Bacteria for Use in Agriculture. Biotechnol. Adv., 16: 729-770.
3. Bishop, J. G., Dean, A. M. and Mitchell-Olds, T. 2000. Rapid Evolution in Plant Chitinases: Molecular Targets of Selection in Plant-pathogen Co-evolution. Proc. Natl. Acad. Sci. USA, 97: 5322–5327.
4. Brick, J. M., Bostock, R. M. and Silverstone, S. E. 1991. Rapid In situ Assay for Indole Acetic Acid Production by Bacteria Immobilized on Nitrocellulose Membrane. Appl. Environ. Microbio., 57: 535–538.
5. Bruehl, G. W. 1987. Soilborne Plant Pathogens. Macmillan, New York.
6. Capper, A. L. and Higgins, K. P. 1993. Application of Pseudomonas fluorescens Isolates to Wheat as Potential Biological Control Agents against Take-all. Plant Pathol., 42: 560–567.
7. Cappuccino, J. C. and Sherman, N. 2010. Microbiology: A Laboratory Manual. Ninth Edition, Cummings Publishing, Benjamin, CA.
8. Cattelan, A. Z., Hartal, P. G. and Fuhrmann, J. J. 1990. Screening for Plant Growth-promoting Rhizobacteria to Promote Early Soybean Growth. Soil Sc. Am. J., 43: 1670–1680.
9. Dijksterhuis, J., Sanders, M., Gorris, L. G. M. and Smid, E. J. 1999. Antibiosis Plays a Role in the Context of Direct Interaction during Antagonism of Paenibacillus polymyxa towards Fusarium oxysporum. J. Appl. Microbiol., 86: 13–21.
10. Duffy, B. K., Ownley, B. H. and Weller, D. M. 1997. Soil Chemical and Physical Properties Associated with Suppression of Take-all of Wheat by Trichoderma koningii. Phytopathol., 87: 1118–1124.
11. El-Bendary, M. A. 2006. Bacillus thuringiensis and Bacillus sphaericus Biopesticides Production. J. Basic Microbiol., 46: 158–170.
12. Emmert, E. A. B. and Handelsman, J. 1999. Biocontrol of Plant Disease: A Gram-positive Perspective. FEMS Microbiol. Lett., 171: 1–9.
13. Felsenstein, J. 1985. Confidence Limits on Phylogenies: An Approach Using the Bootstrap. Evolution, 39: 783-791.
14. Foldes, T., Banhegyi, I., Herpai, Z., Varga, L. and Szigeti, J. 2000. Isolation of Bacillus Strains from the Rhizosphere of Cereals and In vitro Screening for Antagonism against Phytopathogenic, Food-borne Pathogenic and Spoilage Microorganisms. J. Appl. Microbiol., 89: 840–846.
15. Fritze, D. 2004. Taxonomy of the Genus Bacillus and Related Genera: The Serobic Endospore-forming Bacteria. Phytopathol., 94: 1245-1248.
16. Holt, J. G., Krieg, N. R., Sneath, P. H. A., Staley, J. T. and Williams, S. T. 1994. Bergey’s Manual of Determinative Bacteriology. 9th Edition, Williams and Wi1kins, Ba1timore, MD.
17. Jo, H. and Stabb, E. V. 1996. Biocontrol of Soil-borne Plant Pathogens. Plant Cell, 8(10): 1855－1869.
18. Jordan, V., Guilhem, D., Marie, L. B., Bruno, T., Yvan, M. L., Daniel, M., Laurent L, Florence, W. D., and Claire P. C.,, 2013. Plant Growth-promoting Rhizobacteria and Root System Functioning. Frontier. Plant Sci., 4: 356.
19. Jukes, T. H. and Cantor, C. R. 1969. Evolution of Protein Molecules: Mammalian Protein Metabolism. Academic Press, New York, PP. 21-132.
20. Karlidag, H., Esitken, A., Turan, M., and Sahin, F. 2007. Effects of Root Inoculation of Plant Growth Promoting Rhizobacteria (PGPR) on Yield, Growth and Nutrient Element Contents of Leaves of Apple. Sci. Horticult., 114: 16-20.
21. Lee, K. J., Kamala-Kannan, S., Sub, H. S., Seong, C. K. and Lee, G. W. 2008. Biological Control of Phytophthora Blight in Red Pepper (Capsicum annuum L.) Using Bacillus subtilis. World J. Microbiol. Biotechnol., 24: 1139-1145.
22. Lorck, H. 1948. Production of Hydrocyanic Acid by Bacteria. Physiol. Plant, 1: 142–146.
23. Lugtenberg, B. and Kamilova, F. 2009. Plant-growth-promoting Rhizobacteria. Annu. Rev. Microbiol., 63: 541–556.
24. Pal, M. 1998. Diseases of Pulse Crop, Their Relative Importance and Management. J. Mycol. Plant Pathol., 28 (2): 114-122.
25. Palumbo, J. D., Yuen, G. Y., Jochum, C. C., Tatum, K. and Kobayashi, D. Y. 2005. Mutagenesis of β-1,3-glucanase Genes in Lysobacter enzymogenes Strain C3 Results in Reduced Biological Control Activity toward Bipolaris Leaf Spot of Tall Fescue and Pythium Damping-off of Sugar Beet. Phytopathol., 95: 701-707.
26. Pattern, C. L. and Glick, B. R. 1996. Bacterial Biosynthesis of Indole-3-acetic Acid. Can. J. Microbiol., 42: 207–220.
27. Perez, C., Pauli, M. and Bazerque, P. 1990. An Antibacterial Assay by Agar Well Diffusion Method. Acta Bio. Et. Med. Exp., 15: 113-115.
28. Pikovaskya, R. I. 1948. Mobilization of Phosphorus in Soil in Connection with the Vital Activity of Microbial Species. Microbiol., 17: 362–370.
29. Powell, K. A. and Jutsum, A. R. 1993. Technical and Commercial Aspects of Biological Control Products. Pestic. Sci., 37: 315–321.
30. Raaijmakers, J. M., Paulitz, T. C., Steinberg, C., Alabouvette, C. and Moenne-Loccoz, Y. 2009. The Rhizosphere: A Playground and Battlefield for Soilborne Pathogens and Beneficial Microorganisms. Plant Soil, 321: 341–361.
31. Raju, G., Mamta, S., Rameshwar, T. and Suresh, P. 2013. Occurrence and Distribution of Chickpea Diseases in Central and Southern Parts of India. Am. J. Plant Sci., 4: 940-944.
32. Richardson, A. E., Hadobas, P. A., Hayes, J. E., O’Hara, J. E. and Simpson, R. J. 2001. Utilization of Phosphorus by Pasture Plants Supplied with Myoinositol Hexaphosphate Is Enhanced by the Presence of Soil Microorganisms. Plant Soil, 229: 47-56.
33. Prashar, P., Kapoor, N., and Sachdeva, S. 2013. Isolation and Characterization of Bacillus sp with In-vitro Antagonistic Activity against Fusarium oxysporum from Rhizosphere of Tomato. J. Agr. Sci. Tech., 15: 1501-1512
34. Saitou, N. and Nei, M. 1987. The Neighbour-joining Method: A New Method for Reconstructing Phylogenetic Trees. Mol. Biol. Evol., 4: 406-425.
35. Schwyn, B. and Neilands, J. B. 1987. Universal Chemical Assay for the Detection and Determination of Siderophores. Anal. Biochem., 160(1): 47–56.
36. Siddiqui, Z. A., Baghel, G. and Akhtar, M. S. 2007. Biocontrol of Meloidogyne javanica by Rhizobium and Plant Growth-promoting Rhizobacteria on Lentil. World J. Microbiol. Biotechnol., 23: 435–441.
37. Singh, R. K., Kumar, D. P., Solanki, M. K., Singh, P., Srivastva, A. K., Kumar S, Kashyap P. L., Saxena A. K., Singhal P. K., Arora D. K.,, 2013. Optimization of Media Components for Chitinase Production by Chickpea Rhizosphere Associated Lysinibacillus fusiformis B-CM18. J. Basic Microbiol., 53: 451–460.
38. Smibert, R. M. and Krieg N. R. 1994 Phenotypic Characterization. Methods for General and Molecular Biology. American Society for Microbiology, Washington, DC, PP. 607–654.
39. Tamura, K., Peterson, D., Peterson, N., Stecher, G., Nei, M. and Kumar, S. 2011. MEGA5: Molecular Evolutionary Genetics Analysis using Maximum Likelihood, Evolutionary Distance, and Maximum Parsimony Methods. Mol. Biol. Evol., 28: 2731-2739.
40. Teather, R. M. and Wood, P. J. 1982. Use of Congo Red Polysaccharide Interactions in Enumeration and Characterization of Cellulolytic Bacteria from the Bovine Rumen. Appl. Environ. Microbiol., 43: 777–780.
41. Thompson, J. D., Higgins, D. G. and Gibson, T. J. 1994. CLUSTAL W: Improving the Sensitivity of Progressive Multiple Sequence Alignment through Sequence Weighting, Position-specific Gap Penalties and Weight Matrix Choice. Nucleic Acid. Res., 22: 4673-4680.
42. Whipps, J. M. 1997. Ecological Considerations Involved in Commercial Development of Biological Control Agents for Soil-borne Diseases. Modern Soil Microbiology. Marcel Dekker, New York, PP. 525–546.
Journal of Agricultural Science and Technology
Volume 17, Issue 5
September and October 2015
Pages 1365-1377