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Characterization and phylogenetic analysis of halophilic bacteria isolated from rhizosphere soils of coastal plants in Dokdo islands

독도 해안식물로부터 분리된 호염성 세균들의 특성 및 계통학적 분석

  • 유영현 (경북대학교 생명과학부) ;
  • 박종명 (롯데중앙연구소) ;
  • 이명철 (농촌진흥청 국립농업과학원 농업유전자원센터) ;
  • 김종국 (경북대학교 생명과학부)
  • Received : 2015.02.23
  • Accepted : 2015.03.16
  • Published : 2015.03.31

Abstract

To study the halobacterial diversity at the rhizospheric soil of coastal plant native to Dokdo islands, several host plant were selected and its rhizospheric soil was sampled. Soil sample was diluted serially and pure isolation was done by sub-culture using marine agar media. 26 halophilic strains cultivable at the marine medium containig concentration of 9.0% sodium chloride were selected among total 161 isolates. Their partial 16S rRNA gene sequences extracted from genomic DNA were analyzed and partially identified. Furthermore, to identify their genetic relationship, phylogenetic tree was deduced. Total 26 strains were belongs to Firmicutes (30.8%), Gamma proteobacteria (53.8%), Bacteroidetes (7.7%), Alpha proteobacteria (7.7%), and Actinobacteria (7.7%). These results showed the specific difference from previous researches which has been reported the microbial flora of soil or sea water around the Dokdo islands. Furthermore, 4 among 26 halophilic strains grew at above 12.0% NaCl concentrated marine broth, and 2 strains Idiomarina abyssalis LM4H23 and Halomonas huangheensis AS4H13 grew at 15.0% concentration. These halophilic strains thought to overcoming the severe stress like high salt concentration or variation derived from Dokdo-specific climate and might have unknown, specific relationship with their host coastal plant native to Dokdo islands.

독도의 해안에 군락을 이룬 해안식물 근권에서 호염성 및 염내성을 가지는 세균의 분리를 위해 3종의 해안식물의 군락을 선정한 후 각 식물의 군집 하부에서 토양시료를 채취하였다. 시료는 marine broth 한천배지를 이용하여 형태학적인 구분을 통해 순수분리 되었다. 분리된 161개 세균들을 NaCl 9.0% 농도로 조정된 배지에서 생존하는 26개 균주를 선발하여 genomic DNA를 얻은 후, 16S rRNA gene sequence를 증폭하여 부분동정 하였다. 이들의 유연관계 확인을 위해 계통수를 작성한 결과, 이들은 각각 Firmicutes (30.8%), Gamma proteobacteria (53.8%), Bacteroidetes (7.7%), Alpha proteobacteria (7.7%), Actinobacteria (7.7%)에 속하였으며, 이는 기존의 독도 토양 및 해수 미생물상 연구와 특징적 차이를 보인다. 또한, 분리된 세균의 종 조성도 기존 독도 토양 및 해양연구와 유의적으로 상이함을 보였다. 이에 더하여 선발된 26개 균주들 중에서 4균주가 12.0% 이상의 염농도에서 생장하였으며, 이들 중에서 3개 균주가 15.0% 이상의 염농도에서 생장하여 극호염성의 특성을 나타내었으며, 광범위한 염분농도에서도 생장하는 특성을 보였다. 이들은 해안식물 근권에서 독도 특유의 고염분 및 염분변화라는 환경적 스트레스를 극복하며 해안식물과 어떠한 상호작용을 하는 것으로 생각된다.

Keywords

References

  1. Amann, R.I., Ludwig, W., and Schleifer, K.H. 1995. Phylogenetic identification and in situ detection of individual microbial cells without cultivation. Microbiol. Rev. 59, 143-169.
  2. Anan'ina, L.N., Plotnikova, E.G., Gavrish, Elu, Demakov, V.A., and Evtushenko, L.I. 2007. Salinicola socius gen. nov., sp. nov., a moderately halophilic bacterium from a naphthalene-utilizing microbial association. Mikrobiologiia 76, 369-376.
  3. Baati, H., Guermazi, S., Gharsallah, N., Sghir, A., and Ammar, E. 2010. Microbial community of salt crystals processed from Mediterranean seawater based on 16S rRNA analysis. Can. J. Microbiol. 56, 44-51. https://doi.org/10.1139/W09-102
  4. Campbell, B.J., Engel, A.S., Porter, M.L., and Takai, K. 2006. The versatile ${\varepsilon}$-proteobacteria: key players in sulphidic habitats. Nat. Rev. Microbiol. 4, 458-468. https://doi.org/10.1038/nrmicro1414
  5. Cho, B.C. and Azam, F. 1988. Major role of bacteria in biogeochemical fluxes in the ocean's interior. Nature 332, 441-443. https://doi.org/10.1038/332441a0
  6. Choo, Y.S. 2009. Chapter IV. Soil environment of Dokdo island. pp. 108-127, Natural heritage of Korea, Dokdo. Research Institute for Ulleungdo & Dokdo Islands of Kyungpook Nat'l Univ. S.Korea
  7. Cottrell, M.T. and Kirchman, D.L. 2000a. Natural assemblages of marine proteobacteria and members Cytophaga-Flavobacter cluster consuming low-and high-molecular-weight dissolved organic matter. Appl. Environ. Microbiol. 66, 1692-1697. https://doi.org/10.1128/AEM.66.4.1692-1697.2000
  8. Cottrell, M.T. and Kirchman, D.L. 2000b. Community composition of marine bacterioplankton determined by 16S rRNA gene clone libraries and fluorescence in situ hybridization. Appl. Environ. Microbiol. 66, 5116-5122. https://doi.org/10.1128/AEM.66.12.5116-5122.2000
  9. Felsenstein, J. 1985. Confidence limits on phylogeneis: an approach using the bootstrap. Evolution 39, 783-791. https://doi.org/10.2307/2408678
  10. Gonzalez, J.M. and Moran, M.A. 1997. Numerical dominance of a group of marine bacteria in the ${\alpha}$-subclass of the class Proteobacteria in coastal seawater. Appl. Environ. Microbiol. 63, 4237-4242.
  11. Grand, W.D. and Larsen, H. 1989. Extremely halophilic archaebacteria. Order halobacteriales ord. nov., pp. 2216-2233. In Pfennig, N. (ed). Bergey's manual of systematic bacteriology. Vol 3.
  12. Willans and Kilkins. Baltimore. Ham, M.S., Park, Y.M., Sung, H.R., Ryu, C.M., Park, S.H., Sumayo, M., and Ghim, S.Y. 2009. Characterization of Rhizobacteria isolated from Family Solanaceae plants in Dokdo Island. Kor. J. Microbiol. Biotechnol. 37, 110-117.
  13. Ivanova, E.P., Romanenko, L.A., Chun, J., Matte, M.H., Matte, G.R., Mikhailov, V.V., Svetashev, V.I., Huq, A., Maugel, T., and Colwell, R.R. 2000. Idiomarina gen. nov., comprising novel indigenous deep-sea bacteria from the Pacific Ocean, including descriptions of two species, Idiomarina abyssalis sp. nov. and Idiomarina zobellii sp. nov. Int. J. Syst. Evol. Microbiol. 50, 901-907. https://doi.org/10.1099/00207713-50-2-901
  14. Javor, B.J. 1984. Growth potential of halophilic bacteria isolated from solar salt environments: carbon sources and salt requirements. Appl. Environ. Microbiol. 48, 352-360.
  15. Jeon, S.A., Sung, H.R., Park, Y.M., Park, J.H., and Ghim, S.Y. 2009. Analysis of endospore-forming bacteria or nitrogen-fixing bacteria community isolated from plants rhizosphere in Dokdo Island. Kor. J. Microbiol. Biotechnol. 37, 189-196.
  16. Kang, S.M., Radhakrishnan, R., You, Y.H., Joo, G.J., Lee, I.J., Lee, K.E., and Kim J.H. 2014. Phosphate solubilizing Bacillus megaterium mj1212 regulates endogenous plant carbohydrates and amino acids contents to promote mustard plant growth. Indian J. Microbiol. 54, 427-433. https://doi.org/10.1007/s12088-014-0476-6
  17. Kaye, J.Z., Marquez, M.C., Ventosa, A., and Baross, J.A. 2004. Halomonas neptunia sp. nov., Halomonas sulfidaeris sp. nov., Halomonas axialensis sp. nov. and Halomonas hydrothermalis sp. nov.: halophilic bacteria isolated from deep-sea hydrothermalvent environments. Int. J. Syst. Evol. Microbiol. 54, 499-511. https://doi.org/10.1099/ijs.0.02799-0
  18. Kim, B.S., Oh, H.M., Kang, H., Park, S., and Chun, J. 2005. Archaeal diversity in the tidal flat sediment as revealed by 16S rDNA analysis. J. Microbiol. 43, 144-151.
  19. Kim, Y.E., Yoon, H.J., You, Y.H., Kim, H., Seo, Y.G., Kim, M., Woo, J.R., Nam, Y.J., Irina, K., Lee, G.M., et al. 2014. Diversity and characteristics of rhizosphere microorganisms isolated from the soil around the roots of three plants native to the Dokdo Islands. J. Life Sci. 24, 461-466. https://doi.org/10.5352/JLS.2014.24.4.461
  20. Kushner, D.J. 1985. The halobacteriaceae. pp. 171-224. In the Bacteria. Vol. 8. Academic Press. New York, USA.
  21. Lee, S., Ka, J.O., and Song, H.G. 2012. Growth promotion of Xanthium italicum by application of rhizobacterial isolates of Bacillus aryabhattai in microcosm soil. J. Microbiol. 50, 45-49. https://doi.org/10.1007/s12275-012-1415-z
  22. Lee, D.S., Kim, Y.S., Jeong, S.Y., Kang, C.K., and Lee, W.J. 2008. Environmental characteristics and distributions of marine bacteria in the surface sediments of Kamak Bay in winter and summer. J. Environ. Sci. 17, 755-765.
  23. Lee, J.S., Lim, J.M., Lee, K.C., Lee, J.C., Park, Y.H., and Kim, C.J. 2006. Virgibacillus koreensis sp. nov., a novel bacterium from a salt field, and transfer of Virgibacillus picturae to the genus Oceanobacillus as Oceanobacillus picturae comb. nov. with emended descriptions. Int. J. Syst. Evol. Microbiol. 56, 251-257. https://doi.org/10.1099/ijs.0.63734-0
  24. Lim, J.M., Jeon, C.O., and Kim, C.J. 2006. Bacillus taeanensis sp. nov., a halophilic Gram-positive bacterium from a solar saltern in Korea. Int. J. Syst. Evol. Microbiol. 56, 2903-2908. https://doi.org/10.1099/ijs.0.64036-0
  25. Lu, J., Nogi, Y., and Takami, H. 2001. Oceanobacillus iheyensis gen. nov., sp. nov., a deep-sea extremely halotolerant and alkaliphilic species isolated from a depth of 1050 m on the Iheya Ridge. FEMS Microbiol. Lett. 205, 291-297. https://doi.org/10.1111/j.1574-6968.2001.tb10963.x
  26. Maidak, B.L., Cole, J.R., Lilburn, T.G., Parker, C.T., Saxman, P.R., Stredwick, J.M., Garrity, G.M., Li, B., Olsen, G.J., Paranik, S., et al. 2000. The RDP (Ribosomal Database project) continues. Nucleic Acids Res. 28, 173-174. https://doi.org/10.1093/nar/28.1.173
  27. MEthe, B.A., Hiorns, W.D., and Zehr, J.P. 1998. Contrasts between marine and freshwater bacterial community com position: analyses of communities in Lake George and six other Adirondack lakes. Limnol. Oceanogr. 43, 368-374. https://doi.org/10.4319/lo.1998.43.2.0368
  28. Miao, C., Jia, F., Wan, Y., Zhang, W., Lin, M., and Jin, W. 2013. Halomonas huangheensis sp. nov., a moderately halophilic bacterium isolated from a saline-alkali soil. Int. J. Syst. Evol. Microbiol. 64, 915-920.
  29. Na, J.M., Kang, M.S., Kim J.H., Jin, Y.X., Je, J.H., Kim, J.B., Cho, Y.S., Kim, J.H., and Kim, S.Y. 2011. Distribution and identification of halophilic bacteria in solar salts produced during entire manufacturing process. Kor. J. Microbiol. Biotechnol. 39, 133-139.
  30. Park, H.S. and Jeong, M.J. 1996. Isolation and identification of an extremely halophilic bacterium from solar salt. Kor. J. Appl. Microbiol. Biotechnol. 24, 671-677.
  31. Porcel, R., Zamarreno, A.M., Garcia-Mina, J.M., and Aroca, R. 2014. Involvement of plant endogenous ABA in Bacillus megaterium PGPR activity in tomato plants. BMC Plant Biol. 25, 14-36.
  32. Saitou, N. and Nei, M. 1987. The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol. Biol. Evol. 4, 406-425.
  33. Seo, S.T., Kim, K.H., Kim, M.J., Hong, J.S. Park, J.H., and Shin, S.C. 2009. Diversity of fungal endophytes from Pinus koraiensis leaves in Korea. Korean J. Mycol. 37, 108-110. https://doi.org/10.4489/KJM.2009.37.1.108
  34. Seo, Y., Kim, M., You, Y.H., Yoon, J.H., Woo, J.R., Lee, G., and Kim, J.G. 2012. Genetic diversity of endophytic fungi isolated from the roots of halophytes naturally growing in Suncheon Bay. Kor. J. Mycol. 40, 7-10. https://doi.org/10.4489/KJM.2012.40.1.007
  35. Shin, H.T., Park, S.J., Kang, K.H., and Yoo, J.H. 2004. The establishment of conservation area and conservation strategy in Ulnung island (II)-flora and management in Dokdo island, South Korea. Korean J. Environ. Ecol. 18, 221-230.
  36. Siddikee, M.A., Chauhan, P.S., Anandham, R., Han, G.H., and Sa, T. 2010. Isolation, characterization, and use for plant growth promotion under salt stress, of ACC deaminase-producing halotolerant bacteria derived from coastal soil. J. Microbiol. Biotechnol. 20, 1577-1584. https://doi.org/10.4014/jmb.1007.07011
  37. Simbert, R.M. and Kreig, N.R. 1981. General characterization. pp. 409-443. Manual of methods for general bacteriology. American society for Microbiology. Washington, D.C., USA.
  38. Sung, H.R. and Ghim, S.Y. 2010. Bacterial diversity and distribution of cultivable bacteria isolated from Dokdo Island. Kor. J. Microbiol. Biotechnol. 38, 263-272.
  39. VijayAnand, S., Hemapriya, J., Selvin, J., and Kiran, S. 2010. Production and optimization of haloalkaliphilic protease by an extremophile Halobacterium sp. JS1, isolated from thalassohaline environment. Global J. Biotech. Biochem. 5, 44-49.
  40. Woo, P.C., Lau, S.K., Teng, J.L., and Yuen, K.Y. 2008. Then and now: use of 16S rDNA gene sequencing for bacterial identification and discovery of novel bacteria in clinical microbiology laboratories. Clin. Microbiol. Infect. 14, 908-934. https://doi.org/10.1111/j.1469-0691.2008.02070.x
  41. Yoon, J.H., Kang, S.J., Lee, S.Y., Lee, M.H., and Oh, T.K. 2005. Virgibacillus dokdonensis sp. Nov., isolated from a Korean island, Dokdo, located at the edge of the Ease Sea in Korea. Int. J. Syst. Evol. Microbiol. 55, 1833-1837. https://doi.org/10.1099/ijs.0.63613-0
  42. Yoon, J.H., Lee, S.T., and Park, Y.H. 1998. Inter-and intraspecific phylogenetic analysis of the genus Norcardioides and related taxa based on 16S rRNA gene sequences. Int. J. Syst. Bacteriol. 48, 187-194. https://doi.org/10.1099/00207713-48-1-187
  43. You, Y.H., Yoon, H., Kang, S.M., Shin, J.H., Choo, Y.S., Lee, I.J., Lee, J.M., and Kim, J.G. 2012. Fungal diversity and plant growth promotion of endophytic fungi from six halophytes in Suncheon Bay. J. Microbiol. Biotechnol. 22, 1549-1556. https://doi.org/10.4014/jmb.1205.05010
  44. You, Y.H., Yoon, H.J., Kim, H., Lim, S.H., Shin, J.H., Lee, I.J., Choo, Y.S., and Kim, J.G. 2013. Plant growth-promoting activity and genetic diversity of endophytic fungi isolated from native plants in Dokdo Islands for restoration of a coastal ecosystem. J. Life Sci. 23, 95-101. https://doi.org/10.5352/JLS.2013.23.1.95
  45. You, Y.H., Yoon, H.J., Lee, G.S., Woo, J.R., Shin, J.H., Lee, I.J., Rim, S.O., Choo, Y.S., and Kim, J.G. 2011b. Diversity and plant growth-promotion of endophytic fungi isolated from the roots of plants in Dokdo Islands. J. Life Sci. 21, 992-996. https://doi.org/10.5352/JLS.2011.21.7.992
  46. You, Y.H., Yoon, H.J., Woo, J.R., Seo, Y.G., Kim, M., Choo, Y.S., and Kim, J.G. 2011a. Plant growth-promoting activity of endophytic fungi isolated from the roots of native plants in Dokdo Islands. J. Life Sci. 21, 1619-1624. https://doi.org/10.5352/JLS.2011.21.11.1619

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