• Journal of Microbiology and Biotechnology
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• 제18권6호
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• pp.1040-1043
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• 2008

Klebsiella oxytoca CCUG 15788 is resistant to $Ni^{2+}$ at a concentration of 10 mM and grows in an inducible manner when exposed to lower concentrations of $Ni^{2+}$. The complete genomic sequence of a 4.2-kb HindIII-digested fragment of this strain was determined from genomic DNA. It was shown to contain four nickel resistance genes (nirA, nirB, nirC, and nirD) encoding transporter and transmembrane proteins for nickel resistance. When the plasmid pKOHI4, encoding nirABCD, was transformed into Escherichia coli JM109, the cells were able to grow in Tris-buffered mineral medium containing 3 mM nickel. TnphoA'-1 insertion mutants in the four nickel genes nirA, nirB, nirC, and nirD showed nickel sensitivity. The nir genes were heterogeneously expressed in E. coli, suggesting functional roles of these genes in nickel resistance.

• 한국지하수토양환경학회지:지하수토양환경
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• 제19권4호
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• pp.62-69
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• 2014

In the results of monitoring nitrate concentration in more than 8,000 groundwater wells around agro-livestock, the average and maximum nitrate concentration was 9.4 mg/L and 101.2 mg/L, respectively. Since about 31% of the monitoring wells was exceed the quality standard for drinking water, nitrate control such as remediation or source regulation is required to conserve safe-groundwater in South Korea. Typical nitrate-treatment technologies include ion exchange, reverse osmosis, and biological denitrification. Among the treatment methods, biological denitrification by indigenous microorganism has environmental and economic advantages for the complete elimination of nitrate because of lower operating costs compared to other methods. Major mechanism of the process is microbial reduction of nitrate to nitrite and nitrogen gas. Three functional genes (nosZ, nirK, nirS) that encode for the enzyme involved in the pathway. In this work, we tried to develop simple process to determine possibility of natural denitrification reaction by monitoring the functional gene. For the work, the functional genes in nitrate-contaminated groundwater were monitored by using PCR with specific target primers. In the result, functional genes (nosZ and nirK) encoding denitrification enzymes were detected in the groundwater samples. This method can help to determine the possibility of natural-nitrate degradation in target groundwater wells without multiplex experimental process. In addition, for field-remediation application we selected nitrate-contaminated site where 200~600 mg/L of nitrate is continuously detected. To determine the possibility of nitrate-degradation by stimulated-natural attenuation, groundwater was sampled in two different wells of the site and nitrate concentration of the samples was 300 mg/L and 616 mg/L, respectively. Fumarate for different C/N ratio was added into microcosm bottles containing the groundwater to examine denitrification rate depending on carbon concentration. In the result, once 1.5 times more than amount of fumarate stoichiometry required was added, the 616 mg/L of nitrate and 300 mg/L of nitrate were completely degraded in 8 days and 30 days. The nitrite, byproduct of denitrification process, was also completely degraded during the experimental period.

• Environmental Engineering Research
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• 제20권1호
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• pp.51-57
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• 2015

As expected, the expression of denitrifying genes in a Typha wetland (relatively stagnant compared to other ponds), showing higher nitrogen removal efficiency in summer, was affected by temperature. The abundance and gene transcripts of nitrate reductase (narG), nitrite reductase (nirS), nitric oxide reductase (norB), and nitrous oxide reductase (nosZ) genes in seasonal sediment samples taken from the Acorus and Typha ponds of free surface flow constructed wetlands were investigated using quantitative polymerase chain reaction (Q-PCR) and quantitative reverse transcription PCR (Q-RT-PCR). Denitrifying gene copy numbers ($10^5-10^8$ genes $g^{-1}$ sediment) were found to be higher than transcript numbers-($10^3-10^7$ transcripts $g^{-1}$ sediment) of the Acorus and Typha ponds, in both seasons. Transcript numbers of the four functional genes were significantly higher for Typha sediments, in the warm than in the cold season, potentially indicating greater bacterial activity, during the relatively warm season than the cold season. In contrast, copy numbers and expression of denitrifying genes of Acorus did not provide a strong correlation between the different seasons.

• 한국미생물·생명공학회지
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• 제48권1호
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• pp.72-78
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• 2020

The dynamics of nitrogen compounds and RNA-based functional genes were characterized in the nitrification-denitrification coupling process. For the removal of residual ammonium, intermittent aeration was introduced in the denitrification reactor. N₂O production was not observed in both reactors. In both reactors, the nitrifying genes (achaeal-amoA, bacterial-amoA and hor) and denitrifying genes (narG, nirK, norB and nosZ) had a copy number of 3.92 × 10²-7.25 × 10⁵ and 2.85 × 10²-3.06 × 10⁴ per ng of DNA, respectively. These results suggest that denitrification and nitrification reactions occur in both the nitrification and denitrification reactors, respectively. Therefore, the coupling process is a promising one for the conversion of ammonium to nitrogen without generating N₂O.

• 대한환경공학회지
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• 제28권7호
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• pp.752-756
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• 2006

최근 수계의 총질소(T-N) 규제가 강화되면서 기존 BAF 공정의 개선을 위해 독립적인 무산소조가 추가로 도입되었다. 본 연구에 사용된 공정은 유기물과 질산화 중심으로 개발 된 기술인 $Biobead^{(R)}$공법으로 상용화된 상향류의 BAF공정의 하나이다. 독립적인 무산소조의 도입의 타당성을 검토하기 위해 분자생물학적 방법의 하나인 PCR-DGGE기법이 수행되었다. 두 가지 type의 nitrite reductase genes를 통해 진행되었는데, nirS로 암호화된 cytocrome $cd_1$ nitrite reductase gene과 nirK로 암호화된 Cu를 함유한 nitrite reductase gene이다. 이러한 탈질 기능유전자를 이용하여 PCR-DGGE를 통해 탈질 목적으로 순화된 독립적인 무산소조의 탈질미생물의 군집을 해석하였다. PCR 증폭결과, 탈질을 수행하는 무산소조 내에서는 nirS와 nirK유전자 가운데 nirS유전자만 검출되었고, DGGE 분석결과, 최초 식종원으로 이용된 활성슬러지에서는 상대적으로 많은 band들이 검출되는 반면, 무산소조 내에서는 운전일수와 nitrate 부하량이 증가할수록 단일 band로 우점화 하는 경향을 나타내었다. DGGE band에 대한 염기서열 분석결과, 식종 슬러지의 경우 다양한 uncultured bacteria가 나타났으나, nitrate 제거율이 높은 안정화된 무산소조에서는 alcaligenes faecalis 등 특정 탈질미생물이 우점화 되는 것으로 확인되었다. 결론적으로 이러한 탈질미생물 군집특성을 가지는 무산소조의 도입은 96%이상의 안정적인 탈질을 가능하게 하였으며, BAF 공정 개선을 위한 독립적인 무산소조의 도입은 적절한 것으로 판단되었다.

• 한국지하수토양환경학회지:지하수토양환경
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• 제20권7호
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• pp.80-89
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• 2015

Nitrate is on the most seriou pollutant encountered in shallow groundwater aquifer in agricultural area. There are various remediation technologies such as ion exchange, reverse osmosis, and biological denitrification to recover from nitrate contamination. Biological denitrification by indigenous microorganism of the technologies has been reviewed and applied on nitrate contaminated groundwater. In this work, we selected the site where the annual nitrate (NO₃⁻) concentration is over 105 mg/L and evaluated denitrification process with sampled soil and groundwater from 3 monitoring wells (MW4, 5, 6). In the results, the nitrate degradation rate in each well (MW 4, 5, and 6) was 25 NO₃⁻ mg/L/day, 6 NO₃⁻ mg/L/day, and 3.4 NO₃⁻ mg/L/day, respectively. Nitrate degradation rate was higher in batch system treated with 2 times higher fumarate as carbon source than control batch system (0.42M fumrate/1M NO₃⁻), comparing with batch system with soil sample. This result indicates that increase of carbon source is more efficient to enhance denitrification rate than addition of soil sample to increase microbial dynamics. In this work, we also confirmed that monitoring method of functional genes (nirK and nosZ) involved in denitrification process can be applied to evaluated denitrifcation process possibility before application of field process such as in-situ denitrification by push-pull test.

• 한국미생물·생명공학회지
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• 제47권4호
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• pp.630-638
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• 2019

아산화질소는 이산화탄소보다 약 310배 높은 지구온난화 지수를 갖는 주요 온실가스이다. 본 연구에서는 아산화질소 배출저감을 위해 고도처리슬러지를 접종원으로 이용하여 아산화질소 환원 컨소시움을 확보하였다. 이 컨소시움의 우점종은 Sulfurovum (17.95%), Geobacter (14.63%), Rectinema(11.45%)와 Chlorobium (8.24%)이었다. 아산화질소 환원 컨소시움의 활성에 미치는 C/N 비(mol·mol-1), 탄소원의 영향을 조사한 결과, C/N 비 6.3 및 아세트산을 탄소원으로 공급한 조건에서 최대 아산화질소 환원 활성을 나타냈다. 또한, 본 컨소시움의 3,000 ppm 이하의 아산화질소 농도 범위에서 아산화질소 농도가 증가할수록 환원속도도 증가하였다. 속도론적 해석 결과, 아산화질소 환원 컨소시움의 최대 아산화질소 환원 속도는 163.9 ㎍-N·g VSS^-1·h^-1이었다. 본 Consortium은 아산화질소를 N2로 환원하는데 관여를 nosZ 뿐만 아니라, 질산염을 아질산염으로 환원하는 narG, 아질산염을 일산화질소로 환원하는 nirK 유전자 및 일산화질소를 아산화질소를 환원하는 norB 유전자를 모두 보유하고 있었다. 이는 본 컨소시움은 아산화질소 제거 공정 뿐 만 아니라, 탈질공정에도 활용 가능한 유용한 미생물 자원임을 의미한다.