Bacterial Community Profiling during the Manufacturing Process of Traditional Soybean Paste by Pyrosequencing Method

Pyrosequencing을 이용한 전통된장 제조과정 중 세균군집구조의 분석

  • 김용상 (전북대학교 자연과학대학 생명과학과) ;
  • 정도연 ((재)순창발효미생물관리센터) ;
  • 황영태 (우리촌) ;
  • 엄태붕 (전북대학교 자연과학대학 생명과학과)
  • Received : 2011.08.26
  • Accepted : 2011.09.28
  • Published : 2011.09.30


In order to evaluate the diversity and change of bacterial population during the manufacturing process of traditional soybean paste (doenjang), bacterial communities were analyzed using 16S rRNA gene-based pyrosequencing. In rice straw, the most important inoculum source for fermentation, the bacterial sequences with a relative abundance greater than 1% were assigned to four phyla, Proteobacteria (71%), Actinobacteria (20.6%), Bacteroidetes (4.2%), and Firmicutes (1.3%). Unlike bacterial community composition of rice straw, a different pattern of bacterial population in meju was observed with predominantly high abundance (99.1%) of Firmicutes. Phylum composition in young doenjang was almost same as that of meju. Major genera in young doenjang were Bacillus (81.3%), Clostridium (6.9%) and Enterococcus (6.3%) and the predominant species among bacterial population was B. amyloliquefaciens (63.6%). Abundance of the phylum Firmicutes in mature doenjang was 99.98%, which was even higher value than those in meju and young doenjang. Predominant species in mature doenjang were B. amyloliquefaciens (67.3%), B. atrophaeus (12.7%), B. methylotrophicus (6.5%), B. mojavensis (3.2%), and B. subtilis. (2.5%), which were also identified as major species of the microbial flora in meju. These results suggested that rice straw was a primary source for supplement of Bacillus species in manufacturing the traditional doenjang and that some species of Bacillus strains were mainly involved in the fermentation process of traditional doenjang.


Bacillus;bacteria;microbial community;pyrosequencing;soybean paste


Supported by : 농림수산식품기술기획평가원


  1. Ahn, Y.S., Y.S. Kim, and D.H. Shin. 2006. Isolation, identification, and fermentation characteristics of Bacillus sp. with high protease activity from traditional cheonggukjang. Korean J. Food Sci. Technol. 38, 82-87.
  2. Cho, S.J., S.H. Oh, R.D. Pridmore, M.A. Juillerat, and C.H. Lee. 2003. Purification and characterization of proteases from Bacillus amyloliquefaciens isolated from traditional soybean fermentation starter. J. Agric. Food. Chem. 51, 7664-7670.
  3. Choi, K.K., C.B. Cui, S.S. Han, and D.S. Lee. 2003. Isolation, identification and growth characteristics of main strain related to meju fermentation. J. Kor. Soc. Food. Sci. Nutr. 32. 818-824.
  4. Chun. J., J.H. Lee, Y. Jung, M. Kim, S. Kim, B.K. Kim, and Y.W. Lim. 2007. EzTaxon: a web-based tool for the identification of prokaryotes based on 16S ribosomal RNA gene sequences. Int. J. sys. Evol. Microbiol. 57, 2259-2261.
  5. Dischinger, J., M. Josten, C. Szekat, H.Sahl, and G. Bierbaum. 2009. Production of the novel two-peptide lantibiotic lichenicidin by Bacillus licheniformis DSM 13. PLos One 4, 1-11.
  6. Good, I.J. 1953. The population frequencies of species and the estimation of population parameters. Biometrika 40, 237-264.
  7. Grangemard, I., J. Wallach, R. Maget-Dana, and F. Peypoux. 2001. Lichenysin: A more efficient cation chelator than surfactin. Appl. Biochem. Biotechnol. 90, 199-210.
  8. Jeong, J.Y., H.D. Park, K.H. Lee, H.W. Weon, and J.O. Ka. Microbial community analysis and identification of alternative host-specific fecal indicators in fecal and river water samples using pyrosequencing. J. Microbiol. 49, 585-594.
  9. Jung, S.S., J.I. Choi, W.H. Joo, H.H. Suh, A.S. Na, Y.K. Cho, J.Y. Moon, K.C. Ha, D.H. Paik, and D.O. Kang. 2009. Characterization and purification of the bacteriocin produced by Bacillus licheniformis isolated from soybean sauce. J. Life Sci. 19, 994-1002.
  10. Kim, B.S., B.K. Kim, J.H. Lee, M. Kim, Y.M. Lim, and J. Chun. 2008. Rapid phylogenetic dissection of prokaryotic community structure in tidal flat using pyrosequencing. J. Microbiol. 46, 357-363.
  11. Kim, Y.S., M.C. Kim, S.W Kwon, S.J Kim, I.C. Park, J.O. Ka, and H.Y. Weon. 2011. Analysis of bacterial communities in Meju, a Korean traditional fermented soybean bricks, by cultivation-based and pyrosequencing methods. J. Microbiol. 49, 340-348.
  12. Kim, Y.S., S.H. Yun, D.Y. Jeong, K.S. Hahn, and T.B. Uhm. 2010. Isolation of Bacillus licheniformis producing antimicrobial agents against Bacillus cereus and its properties. Kor. J. Microbiol. 46, 270-277.
  13. Kim, T.W., J.H. Lee, M.H. Park, and H.Y. Kim, 2010. Analysis of bacterial and fungal communities in Japanese and Chinese soybean pastes using nested PCR-DGGE. Curr. Microbiol. 60, 315-320.
  14. Kim, T.W., J.H. Lee, S.E. Kim, M.H. Park, H.C. Chang, and H.Y. Kim. 2009. Analysis of microbial communities in doenjang, a Korean fermented soybean paste, using nested PCR-denaturing gradient gel electrophoresis. Int. J. Food Microbiol. 131, 265-271.
  15. Koumoutsi, A., X-H. Chen, J. Vater, and R. Borriss. 2007. DegU and YczE positively regulate the synthesis of Bacillomycin D by Bacillus amyloliquefaciens strain FZB42. Appl. Environ. Microbiol. 73, 6953-6964.
  16. Lane, D.J. 1991. 16S/23S rRNA sequencing, p. 115-148. In E. Stackebrandt and M. Goodfellow (eds.), Nucleic acid techniques in bacterial systematic, John Wiley and Sons, Chister, England.
  17. Li, W. and A. Godzik. 2006. Cd-hit: a fast program for clustering and comparing large sets of protein or nucleotide sequences. Bioinformatics 22, 1658-1659.
  18. Margulies, M., M. Egholm, W.E. Altman, S. Attiya, J.S. Bade, L.A. Bemben et al. 2005. Genome sequencing in microfabricated high-density picoliter reactors. Nature 437, 376-380.
  19. Morohoshi, T., Y. Nakamura, G. Yamazaki, A. Ishida, N.Kato, and T. Ikeda. 2007. The plant pathogen Pantoea ananatis produces N-acylhomoserine lactone and causes center rot disease of onion by quorum sensing. J. Bacteriol. 189, 8333-8338.
  20. Myers, E.W. and W. Miller. 1988. Optimal alignments in linear space. Comput. Appl. Biosci.:CABIOS 4, 11-17.
  21. Parameswaran, P., R. Jalili, L. Tao, S. Shokralla, B. Gharizadeh, M. Ronaghi et al. 2007. A pyrosequencing-tailored nucleotide barcode design unveils opportunities for large-scale sample multiplexing. Nucleic Acids Res. 35, e130.
  22. Park, H.K., B. Gil., and J.K. Kim. 2003. Characteristics of taste components of commercial kochujang. Korean J. Food Sci. Technol. 12, 119-121.
  23. Roh, S.W., K.H. Kim, Y.D. Nam, H.W. Chang, E.J. Park, and J.W. Bae. 2010. Investigation of archaeal and bacterial diversity in fermented seafood using barcoded pyrosequencing. ISME J. 4, 1-16.
  24. Roper, M.C. 2011. Pantoea stewartii subsp. stewartii: lessons learned from a xylem-dwelling pathogen of sweet corn. Mol. Plant Pathol. 12, 628-637.
  25. Ryu, J.C., S.W. Kwon, J.S. Kim, J.S. Suh, B.G. Jung, and S.S. Choi. 2002. Analysis of microbial community structure in soil and crop root system. Korean J. Soil Sci. Fert. 35, 118-126.
  26. Ryu, H.S., M.Y. Shon, S.J. Cho, S.K. Park, and S.W. Lee. 2007. Characterization of antibacterial substance-producing Bacillus subtilis isolated from traditional Doenjang. J. Korean Soc. Appl. Biol. Chem. 50, 87-94.
  27. Schloss, P.D., S.L. Westcott, T. Ryabin, J.R. Hall, M. hartmann, E.B. Hallister, R.A. Lesniewski, B.B. Oakley, D.H. Parks, C.J. Robinson, J.W. Sahl, B. Stres, G.G. Thallinger, D.J. van Horn, and C.F. Weber. 2009. Introducing mothur: open-source, platform-independent, community-supported software for describing and comparing microbial communities. Appl. Environ. Microbiol. 75, 7537-7541.
  28. Scholz, R., K.J. Molohon, J. Nachyigall, J. Vater, A.L. Markley, R.D. Sussmuth, D.A. Mitchell, and R. Borriss. 2011. Plantazolicin, a novel microcin B17/streptolysin S-like natural product from Bacillus amyloliquefaciens FZB42. J. Bacteriol. 193, 215-224.
  29. Secondary planning report for rural-based industry promotion project. 2008. Ministry of Knowledge Economy. Republic of Korea.
  30. Sutyak, K.E., R.E. Wirawan, A.A. Aroutcheva, and M.L. Chikindas. 2008. Isolation of the Bacillus subtilis antimicrobial peptide subtilosin from the dairy product-derived Bacillus amyloliquefaciens. J. Appl Microbiol. 104, 1067-1074.
  31. Yakimov, M.M., K.N. Timmis, V. Wray, and H.H. Fredrickson. 1995. Characterization of a new lipopeptide surfactant produced by thermotolerant and halotolerant subsurface Bacillus licheniformis BAS50. Appl. Environ. Microbiol. 61, 1706-1713.
  32. Yoo, J.Y., H.G. Kim, and W.J. Kim. 1998. Physico-chemical and microbiological changes of traditional meju during fermentation in Kangweondo area. Korean J. Food Sci. Technol. 30, 908-915.