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한반도 주변 해역으로부터 혐기성 미생물의 분리 및 분리 미생물의 특성 분석

Isolation and characterization of anaerobic microbes from marine environments in Korea

  • 김원덕 (한국해양과학기술원 해양생명공학연구센터) ;
  • 이정현 (한국해양과학기술원 해양생명공학연구센터) ;
  • 권개경 (한국해양과학기술원 해양생명공학연구센터)
  • Kim, Wonduck (Marine Biotechnology Research Center, Korea Institute of Ocean Science & Technology) ;
  • Lee, Jung-Hyun (Marine Biotechnology Research Center, Korea Institute of Ocean Science & Technology) ;
  • Kwon, Kae Kyoung (Marine Biotechnology Research Center, Korea Institute of Ocean Science & Technology)
  • 투고 : 2016.02.26
  • 심사 : 2016.04.08
  • 발행 : 2016.06.30

초록

유기산을 생산하는 발효미생물을 획득하기 위해 갯벌, 심해, 염전 등의 퇴적토와 해초시료 등의 시료에 대해 methanogen 배지, acetogen 배지, Clostridium용 배지 등을 이용하여 농후 배양을 실시하였다. 총 8개 시료로부터 65주의 혐기성 미생물을 분리하였으며 이 중 신규성이 높거나 활용성이 높다고 알려진 11종에 대해 계통분석, 성장 양식(growth pattern), 유기산 생산 평가 등을 시도하였다. 분석이 수행된 균주 중 Bacteroidia 강에 속하는 1주 외에는 모두 Clostridia 강에 속하였으며 성장속도는 $1.2h^{-1}$ 이상이었다. 분석이 수행된 7종 중 6종은 아세트산을 생성하였으며, 부가적으로 2균주는 부틸산을, 4균주는 개미산을 생산하였다. 또한 MCWD5 균주는 제공된 포도당의 약 40%를 세포외 고분자물질로 전환시키는 것으로 나타났다. 본 연구를 통하여 국내 연안해역에서 분리된 신규 혐기성 미생물들은 유기산, 고분자 다당류를 생산하는 등 높은 응용성을 지님을 확인할 수 있었다.

Marine bacteria have represented unique physiologies and products which are not discovered from terrestrial organisms. There has been great interest to utilize and develop marine bacteria in many industrial sectors. Recently, we isolated and characterized anaerobic bacteria from various marine environments in Korea to search organic acids fermenting strains. From our enrichment performed under anaerobic condition, 65 strains were isolated and identified by the 16S rRNA gene sequence analysis. Among them, eleven strains were selected for phylogenetical and biochemical analysis. All tested strains were affiliated with Class Clostridia except one with Class Bacteroidia. Most of strains produce acetate (6 strains) with butyrate (2 strains) and/or formate (4 strains). Strain MCWD5 transformed 40% of glucose to extracellular polymeric substances. These results indicate that many novel anaerobic microorganisms which have great potential in commercial application are distributed in the marine environments of Korean Peninsula.

키워드

참고문헌

  1. Andersen, R.J. and Williams, D.E. 2000. Pharmaceuticals from the sea, pp. 55-79. In Hester, R.E. and Harrison, R.M. (eds), Chemistry in the Marine Environment. The Royal Society of Chemistry, Cambridge, UK.
  2. Balch, W.E., Fox, G.E., Magrum, L.J., Woese, C.R., and Wolfe, R.S. 1979. Methanogens: Reevaluation of a unique biological group. Microbiol. Rev. 43, 260-296.
  3. Bowman, K.S., Moe, W.M., Rash, B.A., Bae, H.S., and Rainey, F.A. 2006. Bacterial diversity of an acidic Louisiana groundwater contaminated by dense nonaqueous-phase liquid containing chloroethanes and other solvents. FEMS Microbiol. Ecol. 58, 120-133. https://doi.org/10.1111/j.1574-6941.2006.00146.x
  4. Cavaleiro, A.J., Abreu, A.A., Sousa, D.Z., Pereira, M.A., and Alves, A.A. 2013. The role of marine anaerobic Bacteria and Archaea in bioenergy production, pp. 445-469. In Abdul, M., Elisabeth, G., and Madalena, A. (eds.), Management of Microbial Resources in the Environment. Springer, New York, USA.
  5. Cotta, M.A., Whitehead, T.R., Falsen, E., Moore, E., and Lawson, P.A. 2009. Robinsoniella peoriensis gen. nov., sp. nov., isolated from a swine-manure storage pit and a human clinical source. Int. J. Syst. Evol. Microbiol. 59, 150-155. https://doi.org/10.1099/ijs.0.65676-0
  6. Cotta, M.A., Whitehead, T.R., and Zeltwanger, R.L. 2003. Isolation, characterization and comparison of bacteria from swine faeces and manure storage pits. Environ. Microbiol. 5, 737-745. https://doi.org/10.1046/j.1467-2920.2003.00467.x
  7. Distaso, A. 1912. Contribution a l'etude sur l'intoxication intestinale. Zentralbl. Bakteriol. Parasitenkd. Infektionskr. Hyg. Abt. I. Orig. 62, 433-468.
  8. Fang, M.X., Zhang, W.W., Zhang, Y.Z., Tan, H.Q., Zhang, X.Q., Wu, M., and Zhu, X.F. 2012. Brassicibacter mesophilus gen. nov., sp. nov., a strictly anaerobic bacterium isolated from food industry wastewater. Int. J. Syst. Evol. Microbiol. 62, 3018-3023. https://doi.org/10.1099/ijs.0.034660-0
  9. Faulkner, W. 2002. Marine microbial biodiversity and drug discovery. Abstr. Natural products from marine microorganisms. An international symposium held under the auspices of the European society for marine biotechnology. Greifswald, Germany.
  10. Fenical, W. 1993. Chemical studies of marine bacteria: developing a new resource. Chem. Rev. 93, 1673-1683. https://doi.org/10.1021/cr00021a001
  11. Fontaine, F.E., Peterson, W.H., Mccoy, E., Johnson, M.J., and Ritter, G.J. 1942. A new type of glucose fermentation by Clostridium thermoaceticum n. sp. J. Bacteriol. 43, 701-715.
  12. Gao, Z.M., Xu, X., and Ruan, L.W. 2014. Enrichment and characterization of an anaerobic cellulolytic microbial consortium SQD-1.1 from mangrove soil. Appl. Microbiol. Biotechnol. 98, 465-474. https://doi.org/10.1007/s00253-013-4857-2
  13. Goldstein, E.J. 1995. Anaerobes under assault: from cottage industry to industrialization of medicine and microbiology. Clin. Infect. Dis. 20(Supplement 2), S112-116. https://doi.org/10.1093/clinids/20.Supplement_2.S112
  14. Harris, J.K., Caporaso, J.G., Walker, J.J., Spear, J.R., Gold, N.J., Robertson, C.E., Hugenholtz, P., Goodrich, J., McDonald, D., Knights, D., et al. 2013. Phylogenetic stratigraphy in the Guerrero Negro hypersaline microbial mat. ISME J. 7, 50-60. https://doi.org/10.1038/ismej.2012.79
  15. Himelbloom, B.H. and Canale-Parola, E. 1989. Clostridium methylpentosum sp. nov.: a ring-shaped intestinal bacterium that ferments only methylpentoses and pentoses. Arch. Microbiol. 151, 287-293. https://doi.org/10.1007/BF00406553
  16. Inoue, H., Takimura, O., Kawaguchi, K., Nitoda, T., Fuse, H., Murakami, K., and Yamaoka, Y. 2003. Tin-carbon cleavage of organotin compounds by pyoverdine from Pseudomonas chlororaphis. Appl. Environ. Microbiol. 69, 878-883. https://doi.org/10.1128/AEM.69.2.878-883.2003
  17. Isenbarger, T.A., Finney, M., Rios-Velazquez, C., Handelsman, J., and Ruvkun, G. 2008. Miniprimer PCR, a new lens for viewing the microbial world. Appl. Environ. Microbiol. 74, 840-849. https://doi.org/10.1128/AEM.01933-07
  18. Janssen, P.H. 2004. Propanol as an end product of threonine fermentation. Arch. Microbiol. 182, 482-486. https://doi.org/10.1007/s00203-004-0732-y
  19. Kane, M.D., Brauman, A., and Breznak, J.A. 1991. Clostridium mayombei sp. nov., an $H_2/CO_2$ acetogenic bacterium from the gut of the African soil-feeding termite, Cubitermes speciosus. Arch. Microbiol. 156, 99-104. https://doi.org/10.1007/BF00290980
  20. Kim, S., Jeong, H., and Chun, J. 2007. Clostridium aestuarii sp. nov., from tidal flat sediment. Int. J. Syst. Evol. Microbiol. 57, 1315-1317. https://doi.org/10.1099/ijs.0.64428-0
  21. Kim, S., Jeong, H., Kim, S., and Chun, J. 2006. Clostridium ganghwense sp. nov., isolated from tidal flat sediment. Int. J. Syst. Evol. Microbiol. 56, 691-693. https://doi.org/10.1099/ijs.0.63791-0
  22. Kim, W., Lee, J.H., and Kwon, K.K. 2016. Abyssisolibacter fermentans gen. nov. sp. nov., isolated from deep sub-seafloor sediment. J. Microbiol. 54, 347-352. https://doi.org/10.1007/s12275-016-6048-1
  23. Kotsyurbenko, O.R., Simankova, M.V., Nozhevnikova, A.N., Zhilina, T.N., Bolotina, N.P., Lysenko, A.M., and Osipov, G.A. 1995. New species of psychrophilic acetogens: Acetobacterium bakii sp. nov., A. paludosum sp. nov., A. fimetarium sp. nov. Arch. Microbiol. 163, 29-34. https://doi.org/10.1007/BF00262200
  24. Li, S.L., Whang, L.M., Chao, Y.C., Wang, Y.H., Wang, Y.F., Hsiao, C.J., Tseng, I.C., Bai, M.D., and Cheng, S.S. 2010. Effects of hydraulic retention time on anaerobic hydrogenation performance and microbial ecology of bioreactors fed with glucose-peptone and starch-peptone. Int. J. Hydrogen Energy 35, 61-70. https://doi.org/10.1016/j.ijhydene.2009.10.033
  25. Manivasagan, P. and Kim, S.K. 2014. Extracellular polysaccharides produced by marine bacteria. Adv. Food Nutr. Res. 72, 79-94. https://doi.org/10.1016/B978-0-12-800269-8.00005-1
  26. McCoy, E., Fred, E.B., Peterson, W.H., and Hastings, E.G. 1926. A cultural study of the acetone butyl alcohol organisms. J. Infect. Dis. 39, 457-483. https://doi.org/10.1093/infdis/39.6.457
  27. Meyer, J., Schmidt, A., Michalke, K., and Hensel, R. 2007. Volatilisation of metals and metalloids by the microbial population of an alluvial soil. Syst. Appl. Microbiol. 30, 229-238. https://doi.org/10.1016/j.syapm.2006.05.001
  28. Mohapatra, B.R., Bapuji, M., and Sree, A. 2003. Production of industrial enzymes (amylase, carboxymethylcellulase and protease) by bacteria isolated from marine sedentary organisms. Acta Biotechnol. 23, 75-84. https://doi.org/10.1002/abio.200390011
  29. Pham, V.D., Hnatow, L.L., Zhang, S., Fallon, R.D., Jackson, S.C., Tomb, J.F., DeLong, E.F., and Keeler, S.J. 2009. Characterizing microbial diversity in production water from an Alaskan mesothermic petroleum reservoir with two independent molecular methods. Environ. Microbiol. 11, 176-187. https://doi.org/10.1111/j.1462-2920.2008.01751.x
  30. Raghukumar, C., Vipparty, V., David, J.J., and Chandramohan, D. 2001. Degradation of crude oil by marine cyanobacteria. Appl. Microbiol. Biotechnol. 57, 433-436. https://doi.org/10.1007/s002530100784
  31. Rezgui, R., Ben Ali Gam, Z., Ben Hamed, S., Fardeau, M.L., Cayol, J.L., Maaroufi, A., and Labat, M. 2011. Sporosalibacterium faouarense gen. nov., sp. nov., a moderately halophilic bacterium isolated from oil-contaminated soil. Int. J. Syst. Evol. Microbiol. 61, 99-104. https://doi.org/10.1099/ijs.0.017715-0
  32. Sardessai, Y.N. and Bhosle, S. 2004. Industrial potential of organic solvent tolerant bacteria. Biotechnol. Prog. 20, 655-660. https://doi.org/10.1021/bp0200595
  33. Soriano, S. and Soriano, A. 1948. Nueva bacteria anaerobia productora de una alteracion en sordinas envasadas. Rev. Asoc. Argent. Dietol. 6, 36-41.
  34. Tamura, K., Peterson, D., Peterson, N., Stecher, G., Nei, M., and Kumar, S. 2011. MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol. Biol. Evol. 28, 2731-2739. https://doi.org/10.1093/molbev/msr121
  35. Tracy, B.P., Jones, S.W., Fast, A.G., Indurthi, D.C., and Papoutsakis, E.T. 2012. Clostridia: the importance of their exceptional substrate and metabolite diversity for biofuel and biorefinery applications. Curr. Opin. Biotechnol. 23, 364-381. https://doi.org/10.1016/j.copbio.2011.10.008
  36. Wang, B., Ji, S.Q., Tian, X.X., Qu, L.Y., and Li, F.L. 2015. Brassicibacter thermophilus sp. nov., a thermophilic bacterium isolated from coastal sediment. Int. J. Syst. Evol. Microbiol. 65, 2870-2874. https://doi.org/10.1099/ijs.0.000348
  37. Weusthuis, R.A., Lamot, I., van der Oost, J., and Sanders, J.P. 2011. Microbial production of bulk chemicals: development of anaerobic processes. Trends Biotechnol. 29, 153-158. https://doi.org/10.1016/j.tibtech.2010.12.007
  38. Whitehead, R. 1999. Natural product chemistry. Annu. Rep. Prog. Chem. Sec. B. 95, 183-205. https://doi.org/10.1039/a808577c
  39. Wolfe, R.S. 1999. Anaerobic life-a centennial view. J. Bacteriol. 181, 3317-3320.
  40. Yazdani, S.S. and Gonzalez, R. 2007. Anaerobic fermentation of glycerol: a path to economic viability for the biofuels industry. Curr. Opin. Biotechnol. 18, 213-219. https://doi.org/10.1016/j.copbio.2007.05.002