Gram-positive alkaliphilic microorganisms

Details Gram-positive alkaliphilic microorganisms Gram-positive alkaliphilic microorganisms

1999-01-11

2001-09-18

Lilling, Herbert J. (Department: 1651)

Chemistry: molecular biology and microbiology

Micro-organism, per se ; compositions thereof; proces of...

Bacteria or actinomycetales; media therefor

C435S183000, C435S190000, C435S195000, C435S196000, C435S197000, C435S198000, C435S200000, C435S202000, C435S210000, C435S220000, C435S221000, C435S252310, C435S252400

Reexamination Certificate

active

06291229

ABSTRACT:

The present invention is in the field of microbiology and more particularly in the field of alkaliphilic microorganisms.
BACKGROUND OF THE INVENTION
Alkaliphiles are defined as organisms which exhibit optimum growth in an alkaline pH environment, particularly in excess of pH 8, and generally in the range between pH 9 and 10. Alkaliphiles may also be found living in environments having a pH as high as 12. Obligate alkaliphiles are incapable of growth at neutral pH.
Alkaliphiles may be found in certain everyday environments such as garden soil, presumably due to transient alkaline conditions caused by biological activities including ammonification, sulphate reduction or photosynthesis. A much richer source of a greater variety of alkaliphilic organisms may be found in naturally occurring, stable alkaline environments such as soda lakes.
A more detailed study of soda lakes and alkaliphilic organisms in general is provided in Grant, W. D., Mwatha, W. E. and Jones, B. E. ((1990) FEMS Microbiology Reviews, 75, 255-270), the test of which is hereby incorporated by reference. Lists of alkaline soda lakes may be found in the publications of Grant, W. D. and Tindall, B. J. in
Microbes in Extreme Environments,
(eds. R. A. Herbert and G. A. Codd); Academic Press. London, (1986), pp. 22-54); and Tindall, B. J. in
Halophilic Bacteria,
Volume 1, (ed. F. Rodriquez-Valera); CRC Press Inc., Boca Raton, Fla., (1988), pp. 31-70, both tests are also hereby incorporated by reference.
Alkaliphiles, the majority of which are Bacillus species, have been isolated from non-saline environments and are discussed by Horikoshi, K. and Akiba, T. in
Alkalophilic Microorganisms
(Springer-Verlag, Beriln, Heidelberg, N.Y., (1982). However, alkaliphilic organisms from saline and alkaline environments such as lakes are not discussed therein. Strictly anaerobic bacteria from alkaline, hypersaline, environments have been recently described by Shiba, H. in
Superbugs
(eds. K. Horikoshi and W. D. Grant); Japan Scientific Societies Press, Tokyo and Springer-Verlag, Berlin, Heidelberg, N.Y., (1991), pp. 191-211; and by Nakatsugawa. N., ibid, pp. 212-220.
Soda lakes, which may be found in various locations around the word, are caused by a combination of geological, geographical and climatic conditions. They are characterized by the presence of large amounts of sodium carbonate (or complexes thereof) formed by evaporative concentration, as well as by the corresponding lack of Ca
2+
and Mg
2+
which would remove carbonate ions as insoluble salts. Other salts such as NaCl may also concentrate resulting in environments which are both alkaline and saline.
Despite this apparently harsh environment, soda lakes are nevertheless home to a large population of prokaryotes, a few types of which may dominate as permanent of seasonal blooms. The organisms range from alkaliphilic cyanobacteria to haloalkaliphilic archaeabacteria. Moreover, it is not unusual to find common types of alkaliphilic organisms inhabiting soda lakes in various widely dispersed locations throughout the world such as in the East African Rift Valley, in the western U.S., Tibet, China and Hungary. For example, natronobacteria have been isolated and identified in soda lakes located in China (Wang, D. and Tang, Q., “Natronobacterium from Soda Lakes of China” in
Recent Advances in Microbial Ecology
(Proceedings of the 5th International Symposium on Microbial Ecology, eds. T. Hattori et al.); Japan Scientific Societies Press, Tokyo, (1989), pp. 68-72) and in the western U.S. (Morth, S. and Tindall, B. J. (1985) System. Appl. Microbiol., 6, pp. 247-250). Natronobacteria have also been found in soda lakes located in Tibet (W. D. Grant, unpublished observations) and India (Upasant, V. and Desai, S. (1990) Arch. Microbiol., 154, pp. 589-599).
Alkaliphiles have already made an impact in the application of biotechnology for the manufacture of consumer products. Alkalitolerant enzymes produced by alkaliphilic microorganisms have already found use in industrial processes and have considerable economic potential. For example, these enzymes are currently used in detergent compositions and in leather tanning, and are foreseen to find applications in the food, waste treatment and textile industries. Additionally, alkaliphiles and their enzymes are potentially useful for biotransformations, especially in the synthesis of pure enantiomers. Also, many of the microorganisms described herein are brightly pigmented and are potentially useful for the production of natural colorants.
SUMMARY OF THE INVENTION
The present invention concerns novel aerobic, Gram-positive alkaliphilic bacteria. These bacteria have been isolated from samples of soil, water, segment and a number of other sources, all of which were obtained from in and around alkaline soda lakes. These alkaliphiles have been analyzed according to the principles of numerical taxonomy with respect to each other and also to a collection of known bacteria in order to confirm their novelty. In addition, these bacterial taxa are further circumscribed by an analysis of the lipid components which serve as chemotaxonomic markers.
The present invention also provides data as to the composition of the environments from which the samples containing the microorganisms were obtained, as well as the media required for their efficient isolation and culture such that one of ordinary skill may easily locate such an environment and be able to isolate the organisms of the present invention by following the procedures described herein.
It is also an object of the present invention to provide microorganisms which produce alkalitolerant enzymes. These enzymes are capable of performing their functions at high pH which makes them uniquely suited for applications requiring such extreme conditions. For example, alkalitolerant enzymes may be employed in detergent compositions, in leather tanning and in the food, waste treatment and textile industries, as well as for biotransformations such as the production of pure enantiomers and the production of natural pigments.
The genes encoding these alkalitolerant enzymes may be isolated, cloned and brought to expression in compatible expression hosts to provide a source of larger yields of enzyme products which maybe, if desired, more easily purified and used in various industrial applications, should the wild-type strain fail to produce sufficient amounts of the desired enzyme, or performs poorly under norma, industrial fermentation conditions.


REFERENCES:
patent: 4933287 (1990-06-01), Farin et al.
patent: 5858748 (1999-01-01), Jones et al.
patent: 88/03947 (1988-06-01), None
Collins et al., “Chemotaxonomic Study of an Alkalophilic Bacterium, Exiguobacterium Aurantiacum gen. no., sp. nov.,” J. Gen. Microbiol. (1983) 129:2037-2042.
Durham et al., “Novel Alkaline-and Heat Stable Serine Proteases fromAlkalophilic Bacillussp. Strain GX6638”, J. Bacteriol. (1987) 169(6):2762-2768.
Gee et al., “Properties of a New Group of Alkalophilic Bacteria”, J. Gen. Microbiol. (1980) 117:9-17.
Grant, “Alkaline Environments” in Encyclopedia of Microbiology (1992) vol. 1, pp. 73-80.
Grant, “Alkaliphiles: Ecology, Diversity and Applications”, FEMS Microbiology Reviews (1990) 75:255-270.
Grant, “The Alkaline Saline Environment”, Microbes in Extreme Environment (1986) 1:22-54.
Horikoshi et al., Alkalophilic Microorganisms: A New Microbial World, Japan Scientific Press, Tokyo,, and Springer-Verlag, Berlin (1982) pp. 1-27.
Morth et al., “Variation of Polar Lipid Composition Within Haloalkaliphilic Archaebacteria”, System. Appl. Microbiol. (1985) 6:247-250.
Nakatsugawa, “Novel Methanogenic Archaebacteria Which Grow in Extreme Environments”, Superbugs (K. Horikoshi et al., eds.), Japan Scientific Press, Tokyo and Springer-Verlag, Berlin (1991) Chapter 2, pp. 212-220.
Shiba, “Anaerobic Halophiles”, Superbugs (K. Horikoshi et al., eds.) Japan Scientific Press, Tokyo, and Springer-Verlag, Berlin (1991) Chapter 1, pp. 191-121.
Souza et al., “Characterization of a Novel Extremely Alkalophilic Bacterium”, J. Gen. Microbiol.

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