Journal of Soil and Groundwater Environment (한국지하수토양환경학회지:지하수토양환경)

Korean Society of Soil and Groundwater Environment (한국지하수토양환경학회)
권호 표지

Microbial Adaptation in a Nitrate Removal Column Reactor Using Sulfur-Based Autotrophic Denitrification

질산성 질소 제거를 위한 독립영양 황탈질 칼럼에서의 미생물 적응에 관한 연구

  • Shin, Do-Yun (School of Civil, Urban and Geosystem Engineering, Seoul National University) ;
  • Moon, Hee-Sun (School of Civil, Urban and Geosystem Engineering, Seoul National University) ;
  • Kim, Jae-Young (School of Civil, Urban and Geosystem Engineering, Seoul National University) ;
  • Nam, Kyoung-Phile (School of Civil, Urban and Geosystem Engineering, Seoul National University)
  • 신도연 (서울대학교 지구환경시스템공학부) ;
  • 문희선 (서울대학교 지구환경시스템공학부) ;
  • 김재영 (서울대학교 지구환경시스템공학부) ;
  • 남경필 (서울대학교 지구환경시스템공학부)
  • Published : 2006.04.01
Download PDF
⟨ Previous Next ⟩

Abstract

Two sulfur-based column reactors inoculated with a bacterial consortium containing autotrophic denitrifiers were operated for 100 and 500 days, respectively and nitrate removal efficiency and the adaptation of microbial communities in the columns were monitored with column depths and time. For better understanding the adaptation phenomenon, molecular techniques including 16S rDNA sequencing and DGGE analysis were employed. Although both columns showed about 99% of nitrate removal efficiency heterotrophic denitrifiers such as Cenibacterium arsenioxidans and Geothrix fermentans were found to a significant portion at the initial stage of the 100-day reactor operation. However, as operation time increased, an autotrophic denitrifier Thiobacillus denitrificans became a dominant bacterial species throughout the column. A similar trend was also observed in the 500-day column. In addition, nitrate removal efficiencies were different with column depths and thus bacterial species with different metabolic activities were found at the corresponding depths. Especially, T. denitrificans was successfully adapted and colonized at the bottom parts of the columns where most nitrate was reduced.

본 연구에서는 독립영양 황탈질반응을 이용한 질산성 질소 처리 반응 벽체의 탈질능과 미생물학적 안정성을 확인하기 위하여 황/석회석과 독립영양 황탈질 미생물을 이용한 칼럼 반응기를 상향식으로 500일간 운전하여 시간과 깊이에 따른 질산성 질소의 제거 효율을 분석하였으며, 반응기 내부의 미생물 군집 변화를 16S rDNA-cloning 염기서열 분석법 및 DGGE 기법으로 분석하였다. 실험 결과, 미생물의 대사 활동에 따라 칼럼 깊이 별로 질산성 질소 제거율 및 미생물 군집 분포의 큰 차이가 나타났다. 칼럼 반응기의 질산성 질소 제거율은 99%에 달하였으며, 특히 칼럼 아래쪽에서 질산성 질소 제거율이 매우 높게 나타났다. 시간에 따른 제거율은 칼럼 운전 100일 후부터 큰 차이를 나타내지 않았다. 초기 접종원에서는 독립영양 황탈질 미생물인 OTU DE-1, Thiobacillus denitrificans의 비율이 15%에 불과하였으며 반응기 운전 초기에는 접종원 및 100일 운전 후 반응기의 윗부분에서 종속영양 탈질 미생물인 OTU DE-2, Cenibacterium arsenioxidans와 DE-3, Geothrix fermentans가 78%와 90%로 높은 비율을 차지하여 종속영양탈질 미생물들이 우점종을 차지하였다. 그러나 OTU DE-1은 100일 후에 칼럼 아래쪽에서 94%의 비율을 차지하여 우점종이 되었으며, 500일 운전 후 분석한 결과 칼럼 전체에서 86%를 차지하여 독립영양 황탈질 미생물이 안정적으로 적응하였음을 알 수 있었다.

Keywords

References

  1. 문희선, 장선우, 남경필, 김재영, 2005, 강변여과수의 질산성 질 소 제거를 위한 생물학적 반응벽체의 준 파일럿 실험에 관한 연 구, 대한환경공학회, 27(3), 302-308
  2. Ahn, Y., Park, W., Tatavarty, R., and Kim, I. S., 2004, Comparative analysis of vertical heterogeneity of microbial community in sulfur-packed reactor used for autotrophic nitrate removal, J. Environ. Sci. Health, Part A: Tox./Haz. Subs. Environ. Eng., A 39(7), 1805-1818 https://doi.org/10.1081/ESE-120037879
  3. Altschul, S.F., Gish, W., Miller, W., Myers, E., and Lipman, D. J., 1990, Basic local alignment search tool, J. Mol. Biol., 215, 403-410 https://doi.org/10.1016/S0022-2836(05)80360-2
  4. 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(1), 143-169
  5. Balows, A., Truper, H. G., Dworkin, M., Harder, W., and Schleifer, K-H., 1992, The Prokaryotes, 2nd ed., Springer-Verlag, NY, p. 2638-2657
  6. Batchelor, B. and Lawrence, A.W., 1978, Autotrophic denitrification using elemental sulfur, J. Wat. Poll. Control Federation, 50, 1986-2001
  7. Fan, A.M. and Steinberg, V.E., 1996, Health implication of nitrate and nitrite in drinking water: an update on Methemoglobinemia occurrence and reproductive and developmental toxicity, Regul. Toxicol. Pharmacol., 23, 35-43 https://doi.org/10.1006/rtph.1996.0006
  8. Felsenstein, J., 1985, Confidence limits of phylogenies, an approach using the bootstrap, Evolution, 39(4), 783-791 https://doi.org/10.2307/2408678
  9. Flere, J.M. and Zhang, T.C., 1999, Nitrate removal with sulfurlimestone autotrophic denitrification process, J. Environ. Eng.- ASCE, 125(8), 721-729 https://doi.org/10.1061/(ASCE)0733-9372(1999)125:8(721)
  10. Higgins, D.G. and Sharp, P.M., 1988, CLUSTAL: a package for performing multiple sequence alignment on a microcomputer, Gene, 73, 237-244 https://doi.org/10.1016/0378-1119(88)90330-7
  11. Holt, J.G., Krieg, N.R., Sneath, P.H., Staley, J.T., and Williams, S.T., 1994, Bergey's Manual of Determinative Bacteriology, ninth ed. Williams & Wilkins, Baltimore, p. 361
  12. Hoor, T.T., 1975, A new type of thiosulphate oxidizing, nitrate reducing microorganism: Thiomicrospira denitrificans sp. nov., Neth. J. of Sea Res., 9, 344-351 https://doi.org/10.1016/0077-7579(75)90008-3
  13. Johnson, J.L., 1994, Similarity analysis of rRNAs, In P. Gerhardt, R. G. E. Murray, W. A. Wood, and N. R. Krieg (ed.), Methods for general and molecular bacteriology. American Society for Microbiology, Washington, D.C., p. 683-700
  14. Kelly, D.G. and Wood, A.P., 2000, Confirmation of Thiobacillus denitrificans as a species of the genus Thiobacillus, in the $\beta$-subclass of the Proteobacteria, with strain NCIMB 9548 as the type strain, Int. J. Sys. Evol. Microbiol., 50, 547-550 https://doi.org/10.1099/00207713-50-2-547
  15. Koenig, A. and Liu, L.H., 1996, Autotrophic denitrification of landfill leachate using elemental sulfur, Water Sci. Technol., 34, 469-476
  16. Koenig, A., Zhang, T., Liu, L.H., and Fang, H.H.P., 2005, Microbial community and biochemistry process in autosulfurotrophic denitrifying biofilm, Chemosphere, 58(8), 1041-1047 https://doi.org/10.1016/j.chemosphere.2004.09.040
  17. Korea Ministry of Environment, 2005, 2004년 지하수 수질측 정망 운영결과
  18. Kumar, S., Tomura, K., and Nei, M., 1993, MEGA, Molecular Evolution Genetics Analysis, Version 10. Pennsylvania State University, University Park, PA
  19. Lane, D.J., 1991, 16S/23S rRNA sequencing, In E. Stackebrandt and M. Goodfellow (ed.), Nucleic Acid Techniques in Bacterial Systematics, John Wiley & Sons, Chichester, United Kingdom, p. 115-175
  20. Moon, H.S., Ahn, K.H., Lee, S., Nam, K., and Kim, J.Y., 2004, Use of autotrophic sulfur-oxidizers to remove nitrate from bank filtrate in a permeable reactive barrier system, Environ. Pollut., 129(3), 499-507 https://doi.org/10.1016/j.envpol.2003.11.004
  21. Muyzer, G., De Waal, E.C., and Uitterlinden, A.G., 1993, Profiling of complex microbial populations by denaturing gradient gel electrophoresis analysis of polymerase chain reaction-amplified genes coding for 16S rRNA, Appl. Environ. Microbiol., 59, 695- 700
  22. Saitou, N. and Nei, M., 1987, The neighbor-joining method, a new method for constructing phylogenetic trees, Mol. Biol. Evol., 4(4), 406-425
  23. Stevens, T.O. and McKinley, J.P., 1995, Lithoantotrophic microbial ecosystems in deep basalt aquifers, Science, 270, 450-454 https://doi.org/10.1126/science.270.5235.450
  24. US EPA, 1998, Permeable reactive barrier technnolgies for contaminant remediation, EPA/600/R-98/125
  25. Wagner, M., Amann, R., Lemmer, H., and Schleifer, K.H., 1993, Probing activated sludge with oligonucleotides specific for proteobacteria: Inadequacy of culture-dependent methods for describing microbial community structure, Appl. Environ. Microbiol., 59(5), 1520-1525
  26. Zhang, T.C. and Lampe, D.G., 1999, Sulfur:limestone autotrophic denitrification processes for treatment of nitrate-contaminated water: batch experiments, Water Res., 33(3), 599-608 https://doi.org/10.1016/S0043-1354(98)00281-4