Biodiversity and Phylogenetic Analysis of Streptomyces Collected from Bamboo Forest Soil

대나무 산림토양으로부터 수집한 Streptomyces 속 방선균의 계통학적 다양성

  • Lee, Hyo-Jin (Department of Microbial & Nano Materials, Mokwon University) ;
  • Whang, Kyung-Sook (Department of Microbial & Nano Materials, Mokwon University)
  • 이효진 (목원대학교 미생물나노소재학과) ;
  • 황경숙 (목원대학교 미생물나노소재학과)
  • Received : 2010.08.05
  • Accepted : 2010.09.17
  • Published : 2010.09.30

Abstract

To investigate a quantitative evaluation of the actinobacteria, we have collected samples from various kinds of bamboo forest soil. Each different layers contained $2.7{\times}10^6-2.7{\times}10^8$ CFU/g of actinobacteria which was the highest in litter layers of Sasa boreali forest soil. We obtained 330 actinobacteria from different layers of bamboo forest soil; litter (100 strains), humus (70 strains), and rhizosphere soil (160 strains). Based on the colony morphology (aerial mycelium, substrate mycelium, and soluble pigment), isolates were divided into thirty-six groups and we selected 50 representative isolates. 16S rRNA gene sequence analysis showed Streptomyces was major actinobacteria (94%) and they were categorized as cluster I (2 strains), II (35 strains), III (6 strains), and IV (7 strains), respectively. The diversity index of 50 Streptomyces collected from the bamboo forest soil was calculated with the Shannon-Wiener method. Bamboo litter showed higher diversity index level of 3.33 than that of humus and rhizosphere soil. Also, antibiotic activities of our isolates were investigated against Botrytis cinerea, Xanthomonas campestris, Xanthomonas axonopodis pv. vesicatoria, and Bacillus cereus and found in 74, 16, 25, and 24 strains, respectively.

국내 자생하는 왕대, 분죽, 조릿대, 호마죽과 같은 다양한 대나무 산림토양의 낙엽층, 부식층, 근권토양 내 방선균 밀도 측정 결과, $2.7{\times}10^6-2.7{\times}10^8$ CFU/g로 계수되었으며, 특히 조릿대 낙엽층 내에는 $2.7{\times}10^8$ CFU/g의 매우 높은 밀도로 분포하였다. 본 연구에서는 대나무림 낙엽층으로부터 100균주, 부식층으로부터 70균주 그리고 근권토양으로부터 160균주로 총 330균주의 방선균을 수집하였다. 이들 분리된 균주들의 기균사, 기중균사 및 색소형성 등을 관찰한 결과 36개 방선균 군집으로 분류되었다. 각 그룹으로부터 대표 방선균 50균주를 선발하여 16S rRNA 유전자 염기서열을 해석하고 계통학적 위치를 검토한 결과, 94%가 Streptomyces 속에 속하였으며, cluster I (2균주), II (35균주), III (6균주), 그리고 IV (7균주)에 속하는 특징을 나타내었다. 대나무 산림토양으로부터 수집된 Streptomyces 속 방선균 50균주를 Shannon-Wiener법에 의해 다양성 지수를 산출한 결과, 대나무림 낙엽층으로부터 수집된 방선균의 다양도는 3.33으로 부식층과 근권토양 보다 높게 나타났으며, 근권토양으로부터 수집된 방선균의 88%가 cluster II에 속하는 특징을 나타내었다. 본 연구에서 수집한 방선균을 Botrytis cinerea, Xanthomonas campestris 그리고 Xanthomonas axonopodis pv. vesicatoria에 대해 항균활성능을 검토한 결과, 각 74균주, 16균주, 25균주 그리고 24균주가 항균활성능을 나타내었다.

Keywords

References

  1. Berdy, J. 1974. Recent developments of antibiotic research and classification of antibiotics according to chemical structure. Adv. Appl. Microbiol. 18, 309-406. https://doi.org/10.1016/S0065-2164(08)70573-2
  2. Berdy, J. 1985. Screening, classification and identification of microbial products, pp. 9-31. In M.S. Verral (ed.), Discovery and isolation of microbial products, Society of Chemical Industry, Ellis Horwood Ltd., London, UK.
  3. Berdy, J. 1989. The discovery of new bioactive microbial metabolites: Screening and identification, pp. 3-27. In M.E. Bushell (ed.), Bioactive metabolites from microorganisms, Elsevier, Amsterdam, Netherlands.
  4. Chun, J.S., H.D. Youn, Y.I. Yim, H. Lee, M.Y. Kim, Y.C. Hah, and S.O. Kang. 1997. Streptomyces seoulensis sp. nov. Int. J. Syst. Bacteriol. 47, 492-498. https://doi.org/10.1099/00207713-47-2-492
  5. Dilly, O.J., J. Bloem, A. Vos, and J.C. Munch. 2004. Bacterial diversity in agricultural soils during litter decomposition. Appl. Environ. Microbiol. 70, 468-474. https://doi.org/10.1128/AEM.70.1.468-474.2004
  6. Higton, A.A. and A.D. Roberts. 1998. In B.W. Bycroft (ed.), Dictionary of antibiotics and related substances, pp. 10-18. Chapman and Hall, New York, USA.
  7. Kim, B., A.M. Al-Tal, K.S. Bum, P. Somasundaram, and M. Goodfellow. 2000. Streptomyces thermocoprophilus sp. nov., a cellulase-free endo-xylanase–producing streptomycete. Int. J. Syst. Evol. Microbiol. 50, 505-509. https://doi.org/10.1099/00207713-50-2-505
  8. Kim, B., N. Sahin, D.E. Minnikin, J. Zakrzewska-Czerwinska, M. Mordarski, and M. Goodfellow. 1999. Classification of thermophilic streptomycetes, including the description of Streptomyces thermoalcalitolerans sp. nov. Int. J. Syst. Bacteriol. 49, 7-17. https://doi.org/10.1099/00207713-49-1-7
  9. Kim, H.J., S.C. Lee, and B.K. Hwang. 2006. Streptomyces cheonanensis sp. nov., a novel streptomycete with antifungal activity. Int. J. Syst. Evol. Microbiol. 56, 471-475. https://doi.org/10.1099/ijs.0.63816-0
  10. Kim, S.B. and M. Goodfellow. 2002. Streptomyces thermospinisporus sp. nov., a moderately thermophilic carboxydotrophic streptomycete isolated from soil. Int. J. Syst. Evol. Microbiol. 52, 1225-1228. https://doi.org/10.1099/ijs.0.02003-0
  11. Kim, S.B., C. Falconer, E. Williams, and M. Goodfellow. 1998. Streptomyces thermocarboxydovorans sp. nov. and Streptomyces thermocarboxydus sp. nov., two moderately thermophilic carboxydotrophic species from soil. Int. J. Syst. Bacteriol. 48, 59-68. https://doi.org/10.1099/00207713-48-1-59
  12. Kim, S.B., C.N. Seong, S.J. Jeon, K.S. Bae, and M. Goodfellow. 2004. Taxonomic study of neutrotolerant acidophilic actinomycetes isolated from soil and description of Streptomyces yeochonensis sp. nov. Int. J. Syst. Evol. Microbiol. 54, 211-214. https://doi.org/10.1099/ijs.0.02519-0
  13. Kuster, E. and S.T. Williams. 1964. Selection of media for isolation of Streptomyces. Nature 202, 928-929. https://doi.org/10.1038/202928a0
  14. Lanoot, B., M. Vancanneyt, B. Hoste, K. Vandemeulebroecke, M.C. Cnockaert, P. Dawyndt, Z. Liu, Y. Huang, and J. Swings. 2005. Grouping of streptomycetes using 16S-ITS RFLP fingerprinting. Res. Microbiol. 156, 755-762. https://doi.org/10.1016/j.resmic.2005.01.017
  15. Leadbetter, E.R. 1974. Bergey's manual of determinative bacteriology, 8th ed., pp. 599-881. In R.E. Buchanan and N.E. Gibbons (eds.). The Williams & Wilkins Co., Baltimore, Maryland, USA.
  16. Lechevalier, H.A. 1989. A practical guide to generic identification of actinomycetes. In S.T. Williams, M.E. Sharpe, and J.G. Holt (eds.), Bergey's manual of systematic bacteriology, 4th ed. pp. 2344-2347. The Williams & Wilkins Co., Baltimore, Maryland, USA.
  17. Lee, J.Y., J.Y. Lee, H.W. Jung, and B.K. Hwang. 2005. Streptomyces koyangensis sp. nov., a novel actinomycete that produces 4-phenyl-3-butenoic acid. Int. J. Syst. Evol. Microbiol. 55, 257-262. https://doi.org/10.1099/ijs.0.63168-0
  18. Matsuzawa, Y., O.T. Yoshimoto, H. Naganawa, T. Takeuchi, and H. Umezawa. 1980. Biosynthesis of anthracycline antibiotics by Streptomyces galilaeus. II. structure of new anthracycline antibiotics obtained by microbial glycosidation and biological activity. J. Antibiot. 33, 1341-1347. https://doi.org/10.7164/antibiotics.33.1341
  19. Okazaki, T. 1987. Rare actinomycetes-new breed of actinomycetes. J. Microorganism. 3, 453-461.
  20. Park, D.H., J.S. Kim, S.W. Kwon, C. Wilson, Y.M. Yu, J.H. Hur, and C.K. Lim. 2003. Streptomyces luridiscabiei sp. nov., Streptomyces puniciscabiei sp. nov. and Streptomyces niveiscabiei sp. nov., which cause potato common scab disease in Korea. Int. J. Syst. Evol. Microbiol. 53, 2049-2054. https://doi.org/10.1099/ijs.0.02629-0
  21. Rudi, K., G.H. Kleiberg, R. Heiberg, and J.T. Rosnes. 2007. Rapid identification and classification of bacteria by 16S rDNA restriction fragment melting curve analyses (RFMCA). Food Microbiol. 24, 474-481. https://doi.org/10.1016/j.fm.2006.09.006
  22. Rudi, K., M. Zimonja, and T. N$\ae$s. 2006. Alignment-independent bilinear multivariate modelling (AIBIMM) for global analyses of 16S rRNA gene phylogeny. Int. J. Syst. Evol. Microbiol. 56, 1565-1575. https://doi.org/10.1099/ijs.0.63936-0
  23. Shimazu, A., C.J. Kim, and I.D. Yoo. 1993. Diversity of Actinomycetes Species, morphology and life cycle. Kor. J. Appl. Microbiol. Biotechnol. 21, 88-94.
  24. Stackebrandt, E.W., W. Liesack, and B.M. Goebel. 1993. Bacterial diversity in a soil sample from a subtropical Australian environment as determined by 16S rDNA analysis. FASEB J. 7, 232-236.
  25. Stackebrandt, E.W., W. Liesack, and D. Witt. 1992. Ribosomal RNA and rDNA sequence analyses. Gene 115, 255-260. https://doi.org/10.1016/0378-1119(92)90567-9
  26. Takashi, S. 1987. Cosmopolitan actinomycetes. J. Microorganism 3, 482-492.
  27. Tamura, K., J. Dudley, M. Nei, and S. Kumar. 2007. MEGA4: Molecular evolutionary genetics analysis (MEGA) software version 4.0. Mol. Biol. Evol. 24, 1596-1599. https://doi.org/10.1093/molbev/msm092
  28. Tanaka, Y. and S. Omura. 1990. Metabolism and products of actinomycetes an introduction. Actinomycetol. 4, 13-14. https://doi.org/10.3209/saj.4_13
  29. Thompson, J.D., T.J. Gibson, F. Plewniak, F. Jeanmougin, and D.G. Higgins. 1997. The Clustal_X windows interface: Flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Res. 25, 4876-4882. https://doi.org/10.1093/nar/25.24.4876
  30. Turnidge, J.D. and J.M. Bell. 2005. Antimicrobial susceptibility on solid media, pp. 8-60. In V. Lorian (ed.), Antibiotics in laboratory medicine. The Williams & Wilkins Co., Baltimore, London, UK.
  31. Whang, K.S. and S.H. Yu. 1995. Growth patterns of soil bacteria in different organic nutrient concentrations and isolation of facultative and obligate oligotrophic bacteria. Kor. J. Microbiol. 21, 319-324.
  32. Williams, S.T., M. Goodfellow, G. Alderson, E.M.H. Wellington, P.H.A. Sneath, and M.J. Sackin. 1983. Numerical classification of Streptomyces and related genera. J. Gen. Microbiol. 129, 1743-1813.