Diversity among biosurfactant producing Bacillus spp. from diverse habitats was studied among 77 isolates. Cluster analysis based on phenotypic characteristics using unweighted pair-group method with arithmetic averages (UPGMAs) method was performed. Bacillus isolates possessing high surface tension activity and five reference strains were subjected to amplified 16S rDNA restriction analysis (ARDRA). A correlation between the phenotypic and genotypic characterization of Bacillus spp. is explored. Most of the oil reservoir isolates showing high surface activity clustered with B. licheniformis and B. subtilis, the hot water spring isolates clustered in two ingroups, while the petroleum contaminated soil isolates were randomly distributed in all the three ingroups. Present work revealed that diversity exists in distribution of Bacillus spp. from thermal and hydrocarbon containing habitats where majority of organisms belonged to B. licheniformis and B. subtilis group. Isolate B. licheniformis TT42 produced biosurfactant which reduced the surface tension of water from 72？mNm？1 to 28？mNm？1, and 0.05？mNm？1 interfacial tension against crude oil at 8 0 ° C. This isolate clustered with B. subtilis and B. licheniformis group on the basis of ARDRA. These findings increase the possibility of exploiting the Bacillus spp. from different habitats and their possible use in oil recovery. 1. Introduction Bacillus spp. are main workhorses for biotechnological applications. Their products are in the GRAS list (generally regarded as safe) of US Food and Drug Administration (USFDA) and hence are regarded harmless [1–3]. They produce a variety of products namely. extracellular enzymes, biopolymers, biosurfactants, biopesticides, and so forth from renewable resources and are ecofriendly. Biosurfactants are biologically produced surface-active compounds which are versatile process chemicals, and those from Bacillus spp. additionally possess the property of functionality under extreme conditions of pH, temperature, and salinity [4, 5]. These compounds have property of lowering surface and interfacial tensions of liquids. Biosurfactant production has been reported under thermophilic condition using renewable resources and by using statistically optimized medium [6, 7]. One of the potential uses of biosurfactants is in oil industry with minimum purity specification so that whole-cell broth could be used. The lipopeptide biosurfactants produced by B. subtilis and B. licheniformis are highly potent due to their surface tension reducing ability. B. licheniformis and B. subtilis have
References
[1]
Food Drug Administration, 21 CFR 184:1027, Federal Registration, vol. 48, pp. 329–340, 1983.
[2]
M. Schallmey, A. Singh, and O. P. Ward, “Developments in the use of Bacillus species for industrial production,” Canadian Journal of Microbiology, vol. 50, no. 1, pp. 1–17, 2004.
[3]
E. De Clerck and P. De Vos, “Genotypic diversity among Bacillus licheniformis strains from various sources,” FEMS Microbiology Letters, vol. 231, no. 1, pp. 91–98, 2004.
[4]
I. M. Banat, “Biosurfactants production and possible uses in microbial enhanced oil recovery and oil pollution remediation: a review,” Bioresource Technology, vol. 51, no. 1, pp. 1–12, 1995.
[5]
J. D. Desai and I. M. Banat, “Microbial production of surfactants and their commercial potential,” Microbiology and Molecular Biology Reviews, vol. 61, no. 1, pp. 47–64, 1997.
[6]
S. Joshi, S. Yadav, A. Nerurkar, and A. J. Desai, “Statistical optimization of medium components for the production of biosurfactant by Bacillus licheniformis K51,” Journal of Microbiology and Biotechnology, vol. 17, no. 2, pp. 313–319, 2007.
[7]
S. Joshi, C. Bharucha, S. Jha, S. Yadav, A. Nerurkar, and A. J. Desai, “Biosurfactant production using molasses and whey under thermophilic conditions,” Bioresource Technology, vol. 99, no. 1, pp. 195–199, 2008.
[8]
M. W. Rey, P. Ramaiya, B. A. Nelson, et al., “Complete genome sequence of the industrial bacterium Bacillus licheniformis and comparisons with closely related Bacillus sp.,” Genome Biology, vol. 5, pp. 1–13, 2004.
[9]
E. Z. Ron and E. Rosenberg, “Natural roles of biosurfactants,” Environmental Microbiology, vol. 3, no. 4, pp. 229–236, 2001.
[10]
R. A. Slepecky and H. E. Hemphill, “The genus Bacillus nonmedical,” in The Prokaryotes, A. Ballows, T.G. Trüper, M. Dworkin, and W. Harder, Eds., vol. 2, Springer, New York, NY, USA, 2nd edition, 1992.
[11]
P. G. Carrillo, C. Mardaraz, S. I. Pitta-Alvarez, and A. M. Giulietti, “Isolation and selection of biosurfactant-producing bacteria,” World Journal of Microbiology and Biotechnology, vol. 12, no. 1, pp. 82–84, 1996.
[12]
D. M. Coen, Short Protocols in Molecular Biology, Edited by F. M. Ausubel, R. Brent, R. E. Kingston, D. D. Moore, J. G. Seidman, J. A. Smith, and K. Struhl, Greene Publishing Associates and John Wiley & Sons, New York, 2nd edition, 1992.
[13]
D. D. Moore, Short Protocols in Molecular Biology, Edited by F. M. Ausubel, R. Brent, R. E. Kingston, D. D. Moore, J. G. Seidman, J. A. Smith, and K. Struhl, Greene Publishing Associates and John Wiley & Sons, New York, NY, USA, 2nd edition, 1992.
[14]
H. Suthar, K. Hingurao, A. Desai, and A. Nerurkar, “Evaluation of bioemulsifier mediated Microbial Enhanced Oil Recovery using sand pack column,” Journal of Microbiological Methods, vol. 75, no. 2, pp. 225–230, 2008.
[15]
N. Christofi and I. B. Ivshina, “Microbial surfactants and their use in field studies of soil remediation,” Journal of Applied Microbiology, vol. 93, no. 6, pp. 915–929, 2002.
[16]
N. G. K. Karanth, P. G. Deo, and N. K. Veenanadig, “Microbial production of biosurfactants and their importance,” Current Science, vol. 77, no. 1, pp. 116–126, 1999.
[17]
F. M. Bento, F. A. De Oliveira Camargo, B. C. Okeke, and W. T. Frankenberger, “Diversity of biosurfactant producing microorganisms isolated from soils contaminated with diesel oil,” Microbiological Research, vol. 160, no. 3, pp. 249–255, 2005.
[18]
T. N. Nazina, D. S. Sokolova, A. A. Grigoriyan, Y. F. Xue, S. S. Belyaev, and M. V. Ivanov, “Production of oil-releasing compounds by microorganisms from the Daqing oil field, China,” Mikrobiologiya, vol. 72, no. 2, pp. 206–211, 2003.
[19]
A. A. Bodour, K. P. Drees, and R. M. Maier, “Distribution of biosurfactant-producing bacteria in undisturbed and contaminated arid southwestern soils,” Applied and Environmental Microbiology, vol. 69, no. 6, pp. 3280–3287, 2003.
[20]
B. Austin, D. A. Allen, A. L. Mills, and R. R. Colwell, “Numerical taxonomy of heavy metal-tolerant bacteria isolated from an estuary,” Canadian Journal of Microbiology, vol. 23, no. 10, pp. 1433–1447, 1977.
[21]
B. Austin, J. J. Calomiris, J. D. Walker, and R. R. Colwell, “Numerical taxonomy and ecology of petroleum degrading bacteria,” Applied and Environmental Microbiology, vol. 34, no. 1, pp. 60–68, 1977.
[22]
L. M. Mallory, B. Austin, and R. R. Colwell, “Numerical taxonomy and ecology of oligotrophic bacteria isolated from the estuarine environment,” Canadian Journal of Microbiology, vol. 23, no. 6, pp. 733–750, 1977.
[23]
P. Mavingui, G. Laguerre, O. Berge, and T. Heulin, “Genetic and phenotypic diversity of Bacillus polymyxa in soil and in the wheat rhizosphere,” Applied and Environmental Microbiology, vol. 58, no. 6, pp. 1894–1903, 1992.
[24]
D. Caccamo, T. L. Maugeri, and C. Gugliandolo, “Identification of thermophilic and marine bacilli from shallow thermal vents by restriction analysis of their amplified 16S rDNA,” Journal of Applied Microbiology, vol. 91, no. 3, pp. 520–524, 2001.
[25]
E. M. Jennings and R. S. Tanner, “Biosurfactant producing bacteria found in contaminated and uncontaminated soils,” in Proceedings of the Conference on Hazardous Waste Research, pp. 299–306, 2000.
[26]
M. M. Yakimov, K. N. Timmis, V. Wray, and H. L. Fredrickson, “Characterization of a new lipopeptide surfactant produced by thermotolerant and halotolerant subsurface Bacillus licheniformis BAS50,” Applied and Environmental Microbiology, vol. 61, no. 5, pp. 1706–1713, 1995.
[27]
M. J. McInerney, S. Maudgalya, D. P. Nagle, and R. Knapp, “Critical assessment of the use of microorganisms for oil recovery,” Recent Research in Developmental Microbiology, vol. 6, pp. 269–284, 2002.
[28]
M. M. Yakimov, M. M. Amro, M. Bock et al., “The potential of Bacillus licheniformis strains for in situ enhanced oil recovery,” Journal of Petroleum Science and Engineering, vol. 18, no. 1-2, pp. 147–160, 1997.
[29]
M. Javaheri, G. E. Jenneman, M. J. McInerney, and R. M. Knapp, “Anaerobic production of a biosurfactant by Bacillus licheniformis JF-2,” Applied and Environmental Microbiology, vol. 50, no. 3, pp. 698–700, 1985.
[30]
A. S. Nerurkar, “Structural and molecular characteristics of Lichenysin and its relationship with surface activity,” in Biosurfactants, R. Sen, Ed., pp. 304–315, Landes Bioscience and Springer Science + Business Media, N.Y. USA., 2010.
