Phenotypic and Genotypic Characterization of facultative and obligate Alkalophilic Bacillus sp. isolated from Saudi Arabia alkaline soils
Keywords:Bacillus cohnii, 16S rRNA, obligate alkalophiles, facultative alkalophiles, Al-qunfotha city, Saudi Arabia
Isolation and identification of new alkalophilic Bacillus strains have been increasing interest due to their possessing valuable and commercially interesting enzymes. In the present study, a total six obligate and facultative alkalophilic isolates were isolated from desert soil around the Al-qunfotha city, Saudi Arabia. All isolates were phenotypically and genotypically characterized. Among these isolates; AS3, AS4, AS5 and AS6 could grow at pH 9, 10, 11 and 12, but could not grow at pH 7, indicates that these isolates are obligate alkalophiles. While, isolates AS1 and AS2 grew at pH ranged from 7 to 10, but could not grow at pH 11 and 12, suggesting that they could be facultative alkalophiles. All isolates could hydrolyze casein and starch, indicating that they possess interesting amylase and protease enzymes. Based on 16S rDNA data, the phylogenetic analysis of these strains showed that all six alkalophilic Bacillus belonged to Bacillus cohnii with 99% similarity. The nucleotide sequences of 16S rRNA gene for the six isolates were deposited in a Gene-bank under accession numbers; KP053301, KP053302, KP053303, KP053304, KP053305 and KP053306, respectively.
. Assaeedi, A., and Osman, G. (2012). Isolation and characterization of Gram negative obligate and facultative alkalophilic Bacillus sp. from desert soil of Saudi Arabia. African Journal of Biotechnology, 11:9816-9820.
. van Berkum, P., Beyene, D., Eardly, B.D. (1996). Phylogenetic relationships among Rhizobium species nodulating the common bean (Phaseolus vulgaris L.). Int. J. Syst. Bacteriol., 46: 240-244.
. Claus, D. and Berkeley, R.C.W. (1986). Genus Bacillus Cohn, 1872. In: Sneath, P.H.A., Mair, N.S., Sharpe, M.E. and Holt. J.G., Eds., Bergey’s Manual of Systematic Bacteriology, The Williams & Wilkins Co., Baltimore, 2, pp. 1105-1139.
. Elbanna, K., Elbadry, M. and Gamal-Eldin, H. (2009). Genotypic and phenotypic characterization of rhizobia that nodulate Snap bean (Phaseolus vulgaris L.) in Egyptian soils. Syst. Appl. Microbiol., 32, 522–530.
. Elbanna, K., Elnaggar, S. and Bakeer, A. (2014). Characterization of Bacillus altitudinis as a New Causative Agent of Bacterial Soft Rot. Journal of Phytopathology (in press).
. Fritze, D., Flossdorf, J., Claus, D. (1990). Taxonomy of alkaliphilic Bacillus strains. Int. J. Syst. Bacteriol., 40: 92-97.
. Grant, W.D. (1988). Bacteria from alkaline, saline environments and their potential in biotechnology. J. Chem. Technol. Biotechnol., 42:291–94.
. Grant, W.D., Tindall, B.J. (1986). The alkaline saline environment. In: Herbert, R.A., Codd, G.A. (eds). Microbes in Extreme Environments. Academic Press, London, pp 22–54.
. Grant, W.D., Mwatha, W.E., Jones, B.E. (1990). Alkaliphiles: Ecology, diversity and applications. FEMS Microbiol. Rev., 75:255–270.
. Guffanti, A.A., Finkelthal, O., Hicks, D.B., Falk, L., Sidhu, A., Garro, A., Krulwich, T.A. (1986). Isolation and characterization of new Facultatively Alkalophilic strains of Bacillus species. J. Bacteriol., 167: 766-733.
. Horikoshi, K., Akiba, T. (1982). Alkalophilic Microorganisms, a New Microbial World. Japan Scientific Societies Press and Springer Verlag.
. Horikoshi, K. (1999). Alkaliphiles: some application of their products for biotechnology. Microbiol. Mol. Biol. Rev., 63: 735-750.
. Ito, S., Kobayashi, T., Ara, K., Ozaki, K., Kawai, S., Hatada, Y. (1998). Alkaline detergent enzymes from alkaliphiles: enzymatic properties, genetics, and structures. Extremophiles, 2: 185-190.
. Joung, K.B., Cote, J.C. (2002). Evaluation of ribosomal RNA gene restriction patterns for the classification of Bacillus species and related genera. J. Appl. Microbiol., 92: 97-108.
. Krulwich, T.A. (1986). Bioenergetics of alkalophilic bacteria. J. Membr. Biol., 89:113–25.
. Krulwich, T.A., Guffanti, A.A. (1983). Physiology of acidophilic and alkalophilic bacteria. Adv. Microb. Physiol., 24:173–214.
. Krulwich, T.A., Guffanti, A.A. (1989). Alkalophilic bacteria. Ann. Rev. Microbiol., 43:435–63.
. Krulwich, T.A., Guffanti, A.A., Seto-Young, D. (1990). pH homeostasis and bioenergetic work in alkalophiles. FEMS Microbiol. Rev., 6:271–78.
. Kumar, C.G., Tiwari, M.P., Jany, K.D. (1997). Screening and isolation of alkaline protease producers from soda soils of Karnal, India. Proceedings of First National Symposium on Extremophiles, March 20–21, Hamburg, Germany (Abstract no. PE071).
. Marchesi, J.R., Sato, T., Weightman, A.J., Martin, T.A., Fry, J.C., Hiom, S.J., Dymock, D., Wade, W.G. (1998). Design and evaluation of useful bacterium-specific PCR primers that amplify genes coding for bacterial 16S rRNA. Appl. Environ. Microbiol., 64: 795-799.
. Guerra-Cantera, M.A.R.V. & Raymundo, A.K. (2005). Utilization of a polyphasic approach in the taxonomic reassessment of antibiotic- and enzyme-producing Bacillus sp. isolated from the Philippines. World J. Microbiol. Biotechnol., 21: 635-644.
. Martins, R.F., Davids, W., Al-Soud, W.A., Levander, F., Rådström, P. and Hatti-Kaul, R. (2001). Starch-hydrolyzing bacteria from Ethiopian soda lakes. Extremophiles, 5: 135–144.
. Muntyan, M.S., Popova, I.V., Bloch, D.A., Skripnikova, E.V., Ustiyan, V.S. (2005). Energetics of alkalophilic representatives of the genus Bacillus. Biochemistry, 70: 137-142.
. Nielsen, P., Fritze, D. and Priest, F.G. (1995). Phenetic diversity of alkaliphilic Bacillus strains: proposal for nine new species. Microbiology, 141: 1745–1761.
. Saitou, N., Nei, M. (1987). The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol. Biol. Evol., 4: 406-425.
. Salama, A.M., Aggab, A.M., Ramadani, A.S. (1993). Alkalophily among microorganisms inhabiting virgin and cultivated soils along Makkah-Al-Taif road, Saudi Arabia. J. King Saud Univ.: Sci., 5: 69-85.
. Sanger, F., Nicklen, S., Coulson, A.R. (1977). DNA sequencing with chain-terminating inhibitors. Proc. Natl. Acad. Sci. USA., 74: 5463-5467.
. Schallmey, M., Singh, A., Ward, O.P. (2004). Developments in the use of Bacillus species for industrial production. Can. J. Microbiol., 50: 1-17.
. Takami, H. and Horikoshi, K. (2000). Analysis of the genome of an alkaliphilic Bacillus strain from an industrial point of view. Extremophiles, 4, 99–108.
. Tindall, B.J., Mills, A.A., Grant, W.D. (1980). An alkalophilic red halophilic bacterium with a low magnesium requirement from a Kenyan soil lake. J. Gen. Microbiol., 116:257–60.
. Wang, S.Y., Wu, S.J., Thottappilly, G., Locy, R.D., Singh, N.K. (2001). Molecular cloning and structural analysis of the gene encoding Bacillus cereus exochitinase chi36. J. Biosci. Bioeng. 92: 59-66.
. Woese, C.R. (1987). Bacterial Evolution. Microbiol. Rev., 51: 221-271.
. Xu, D., Cote, J. (2003). Phylogenetic relationships between Bacillus species and related genera inferred from comparison of 3' end 16S rDNA and 5' end 16S-23S ITS nucleotide sequences. Int. J. Syst. Evol. Microbiol., 53: 695-704.