Dynamics of Phylogenetic Diversity and its influence on the production of Extracellular Protease by moderately Halotolerant Alkaliphilic Bacteria Acinetobacter baumannii GTCR407 Nov.

  • Thiyagarajan Gurunathan Department of Biotechnology, Central Leather Research Institute, Adyar, Chennai-600 020, Tamilnadu, India.
  • S. Gowtham Kumar Genetics Division, Central Research Laboratory, Chettinad University, Kelambakkam, Chennai-603103, India.
  • C. Anbu Selvam Department of Pharmacology, Sri Manakula Vinayagar Medical College and Hospital, Pondicherry-605107, India.
  • Asit Baran Mandal Chemical Laboratory, Central Leather Research Institute, Adyar, Chennai-600 020, Tamilnadu, India.
Keywords: Haloalkaliphilic, Extracellular protease, Phylogenetic diversity, Microbial population dynamics, Acinetobacter baumannii

Abstract

New characters emerge in the population of microorganisms living in the extreme environments due to its adaptation to ecological association. The microorganisms living in saline habitat utilize complex nutrients by adopting different strategies in Deoxyribonucleic Acid (DNA) and Ribonucleic Acid (RNA), which are related to their metabolic and ecological diversities. Isolation and characterization of the organisms producing extracellular protease from such environment were the prime focus of this investigation, which can indicate the importance of metabolic diversity in phylogeny. Norberg medium was used to isolate halotolerant microorganisms from salt-cured skin. The isolates were screened for high activity of protease and the strain showing maximum activity of protease was taken for further studies. The biochemical characterization and 16s ribosomal RNA sequencing studies confirm that the isolate is Acinetobacter baumannii. Moreover, hydrolysis positive for starch and casein, negative for gelatin shows that the organism is a variant form of A. baumannii. Cell growth parameters such as pH and temperature were optimized and their values are 8 and 37oC respectively. The extracellular production of protease was optimized in the suitable medium and its enzyme activity was 165μg/ml/min. The results imply that the isolate had acquired operational genes through lateral gene transfer (LGT) probably from unrelated species in the environment. This indicates that the isolate identified possesses metabolic and ecological diversities with values of phylogenetic delineation.

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References

[1]. Towner, K. (2006). The Genus Acinetobacter. Prokaryotes, 6:746-758.
[2]. Schreckenberger, P.C., Daneshvar, M.I., Weyant, R.S. & Hollis, D.G. (2003). Acinetobacter, Achromobacter, Chryseobacterium, Moraxella and other non-fermentative Gram-negative rods. In: Murray P.R et al., Eds., Manual of Clinical Microbiology. 8th Ed, ASM Press, Washington, DC, pp. 749-779.
[3]. Bergogne Berezin, E. & Towner, K.J. (1996). Acinetobacter sp. as nosocomial pathogens: microbiological, clinical, and epidemiological features. Clin. Microbiol. Rev., (9): 148-165.
[4]. Gerner smidt, P. & Tjernberg, I. (1993). Acinetobacter in Denmark. II: Molecular studies of the Acinetobacter calcoaceticus-Acinetobacter baumannii complex. Acta. Pathol. Microbiol. Immunol. Scand., (101): 826-832.
[5]. Houang, E.T.S., Chu, Y.W., Chu, K.Y., Ng, K.C., Leung, C.M. & Cheng, A.F.B. (2003). Significance of genomic DNA group delineation in comparative studies of antimicrobial susceptibility of Acinetobacter spp. Antimicrob. Agents Chemother., (47): 1472-1475.
[6]. Spence, R.P., Towner, K.J., Henwood, C.J., James, D., Woodford, N. & Livermore, D.M. (2002). Population structure and antibiotic resistance of Acinetobacter DNA group 2 and 13TU isolates from hospitals in the UK. J. Med. Microbiol., (51): 1107-1112.
[7]. Nemec, A., Dijkshoorn, L. & Jezek, P. (2000). Recognition of two novel phenons of the genus Acinetobacter among non-glucose-acidifying isolates from human specimens. J. Clin. Microbiol., (38): 3937-3941.
[8]. Juni, E. (1984). Genus III: Acinetobacter Brisou et Prevot 1954. In: Krieg N.R., Holt J.G., Eds., Bergy’s Manual of Systematic Bacteriology. Williams and Wilkins, Baltimore, MD, pp. 303-307.
[9]. Rossau, R., Vanlandschoot, A., Gillis, M. & Deley, J. (1991). Taxonomy of Moraxellacea fam.nov., a new bacterial family to accommodate the genera Moraxella, Acinetobacter, and Psychrobacter and related organisms. Int. J. Syst. Bacteriol., (41): 310-319.
[10]. Rossau, R., Vandenbussche, G., Thielemans, S., Segers, P., Grosch, H., Gothe, E., Mannheim, W. & Deley, J. (1989). Ribosomal ribonucleic acid cistron similarities and deoxyribonucleic acid homologies of Neisseria, Kingella, Eikenella, Alysiella and centers for disease control groups EF-4 and M-5 in the emended family Neisseriaceae. Int. J. Syst. Bacteriol., (39): 185-198.
[11]. Van Landschoot, A., Rossau, R. & De Ley, J. (1986). Intra- and intergeneric similarities of the ribosomal ribonucleic acid cistrons of Acinetobacter. Int. J. Syst. Bacteriol., 36(2):150-160.
[12]. Johnson, J.L., Anderson, R.S. & Ordal, E.J. (1970). Nucleic acid homologies among oxidase negative Moraxella species. J. Bacteriol., (101): 568-573.
[13]. Baumann, P., Doudorof, M. & Stanier, R.Y. (1968). A study of the Moraxella group II: Oxidative negative species (genus Acinetobacter). J. Bacteriol., (95): 1520-1541.
[14]. Allers, T. & Mevarech, M. (2005). Archeal genetics-the third way. Nature Rev. Genet., (6): 58-73.
[15]. Norberg, P. & Vonhofst. B. (1969). Proteolytic enzymes from extremely halophilic bacteria. J. Gen. Microbiol., (55): 251-256.
[16]. Wikstrom, M., Elwing, H. & Linde, A. (1981). Determination of proteolytic activity: A sensitive and simple assay utilizing substrate adsorbed to a plastic surface and radial diffusion in gel. Anal. Biochem., (118):240-246.
[17]. Hagihara, B. (1958). The Enzymes. Vol., 4., Academic Press, Inc., New York.
[18]. Lowry, O.H., Rosebrough, N.J., Farr, A.L. & Randall, R.J. (1951). Protein measurement with Folin phenol reagent. J. Biol. Chem., (193): 265-275.
[19]. Ronald, M.A. (2005). Handbook of media for environmental microbiology. 2nd edition. CRC Press, ISBN 0849335604, 9780849335600.
[20]. Nicholson, C.A. & Fathepure, B.Z. (2004). Biodegradation of Benzene by Halophilic and Halotolerant Bacteria under Aerobic Conditions. Appl. Environ. Microbiol., (70): 1222-1225.
[21]. Zhang, W.Z., Xue, Y.F., Ma, Y.H., Zhou, P.J., Ventosa, A. & Grant, W.D. (2002). Salinicoccus alkaliphilus sp. Nov.: A novel alkaliphile and moderate halophile from Baer Soda Lake in inner Mongolia Autonomous Region, China. Int. J. Syst. Evol. Microbiol., (52): 789-793.
[22]. Studdert, C.A., Seitz, M.K.H., Gil, M.I.P., Sanchez, J.J. & de Castro, R.E. (2001). Purification and biochemical characterization of the Haloalkaliphilic archeon Natronococcus occultus extracellular serine protease. J. Basic Microbiol., (41): 375-383.
[23]. Kushner, D.J. (1993). Growth and nutrition of halophilic bacteria. In: Vreeland R.H., and Hochstein L.I., Eds., The biology of halophilic bacteria. CRC Press, Inc., Boca Raton, Fla, pp. 87-103.
[24]. Uchiyama, T., Abe, T., Ikemura, T. & Watanabe, K. (2005). Substrate-induced gene-expression screening of environmental metagenome libraries for isolation of catabolic genes. Nature Biotechnol., (23): 88-93.
[25]. Handelsman, J. (2004). Metagenomics: Application of genomics to uncultured microorganism. Rev. Microbiol Mol. Biol., (68): 669-685.
[26]. Quesada E., Ventosa A., Rodriguez-Valera F., Megias L. & Ramos-Cormenzana A. (1983). Numerical taxonomy of moderately halophilic Gram negative bacteria from hyper saline soils. J. Gen. Microbiol., (129): 2649-2657.
[27]. Oren, A. (2002). Diversity of halophilic microorganisms: Environments, phylogeny, physiology, and applications. J. Ind. Microbiol. Biotechnol., (28): 56-63.
[28]. Lio, P. (2002). Investigating the relationship between genome Structure, Composition, and Ecology in Prokaryotes. Mol. Bio. Evol., (19): 789-800.
[29]. Ochman, H., Lawrence, J.G. & Groisman, E.A. (2000). Lateral gene transfer and the nature of bacterial innovation. Nature, (405): 299-304.
[30]. Karlin, S., Campbell, A.M. & Mrazek, J. (1998). Comparative DNA analysis across diverse genomes. Ann. Rev. Genet., (32): 185-225.
[31]. Jain, R., Rivera, M.C. & Lake, J.A. (1999). Horizontal gene transfer among genomes: The complexity hypothesis. Proc. Natl. Acad. Sci. USA., (96): 3801-3806.
[32]. Rosenshine, I., Tchelet, R. & Mevarech, M. (1989). The mechanism of DNA transfer in the mating system of an archaebacterium. Science, (245): 1387-1389.
[33]. Cryz, S.J. & Iglewski, B.H. (1980). Production of Alkaline protease by Pseudomonas aeruginosa. J. Clin. Microbiol., (12): 131-133.
[34]. Morihara, K. (1964) Production of elastase and proteinase by Pseudomonas aeruginosa. J. Bacteriol., (88): 745-757.
[35]. Gavigan, J.A., Marshall, L.M. & Dobson, A.D.W. (1999). Regulation of polyphosphate kinase gene expression in Acinetobacter baumannii 252. J. Microbiol: Genet. Mol. Biol., (145): 2931-2937.
[36]. Horikoshi, K. (1999). Alkaliphiles: Some application of their products for biotechnology. Microbiol. Mol. Biol. Rev., (63): 735-750.
[37]. Prakasham, R.S., Rao, C.S. & Sarma, P.N. (2006). Green gram husks an inexpensive substrate for alkaline protease production by Bacillus sp. in solid state fermentation. Bio Res. Technol., (97): 1449-1454.
[38]. Krulwich, T.A., Ito, M., Hicks, D.B., Gilmour, R. & Guffanti, A.A. (1998). pH Homeostasis and ATP synthesis: Studies of two processes that necessitate inward proton translocation in extremely alkaliphilic Bacillus sp. Extremophiles, (2): 217-222.
[39]. Ray, M.K., Devi, K.U., Kumar, G.S. & Shivaji, S. (1992). Extracellular protease from the Antarctic yeast Candida humicola. Appl. Env. Microbiol., (58): 1918-1923.
[40]. Votruba, J., Pazlarova, J., Dvorakova, M., Vachova, L., Strnadova, M., Kucerova, H., Vinter, V., Zourabian, R. & Chaloupka, J. (1991). External factors involved in the regulation of synthesis of an extracellular proteinase in Bacillus megaterium: effect of temperature. Appl. Microbiol. Biotechnol., (35): 352-357.
[41]. Frankena, J., Koningstein, G.M., Van- Verseveld, H.W. & Stouthamer, A.H. (1986). Effect of different limitations in chemostat cultures on growth and production of exocellular protease by Bacillus licheniformis. Appl. Microbiol. Biotechnol., (24): 106-112.
Published
2010-07-01
How to Cite
Gurunathan, T., Kumar, S., Selvam, C., & Mandal, A. (2010). Dynamics of Phylogenetic Diversity and its influence on the production of Extracellular Protease by moderately Halotolerant Alkaliphilic Bacteria Acinetobacter baumannii GTCR407 Nov. Journal of Advanced Laboratory Research in Biology, 1(1), 23-30. Retrieved from https://e-journal.sospublication.co.in/index.php/jalrb/article/view/9
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