• 한국물환경학회지
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• 제36권3호
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• pp.220-228
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• 2020

Anaerobic ammonium oxidation (AMX) is a cost-efficient biological nitrogen removal process. The coexistence of ammonia-oxidizing bacteria (AOB) in an AMX reactor is an interesting research topic as a nitrogen-related bacterial consortium. In this study, a sequencing batch reactor for AMX (AMX-SBR) was operated with a conventional activated sludge. The AOB in an AMX bioreactor were identified and quantified using terminal restriction fragment length polymorphism (T-RFLP) and real-time qPCR. A T-RFLP assay based on the ammonia monooxygenase subunit A (amoA) gene sequences showed the presence of Nitrosomonas europaea-like AOB in the AMX-SBR. A phylogenetic tree based on the sequenced amoA gene showed that AOB were affiliated with the Nitrosomonas europaea/mobilis cluster. Throughout the enrichment period, the AOB population was stable with predominant Nitrosomonas europaea-like AOB. Two OTUs of amoA_SBR_JJY_20 (FJ577843) and amoA_SBR_JJY_9 (FJ577849) are similar to the clones from AMX-related environments. Real-time qPCR was used to quantify AOB populations over time. Interestingly, the exponential growth of AOB populations was observed during the substrate inhibition of the AMX bacteria. The specific growth rate of AOB under anaerobic conditions was only 0.111 d^-1. The growth property of Nitrosomonas europaea-like AOB may provide fundamental information about the metabolic relationship between the AMX bacteria and AOB.

• Biotechnology and Bioprocess Engineering:BBE
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• 제2권2호
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• pp.77-81
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• 1997

Removal of nitrogen compound from waste water is essential and often accomplished by biological process. To prevent washout and to develop an efficient bioreactor, immobilization of sutibal microorganisms could be sensible approach. Strains and permeabilized cell encapsulated in cellulose nitrate microcapsules and immobilized on polystyrene films were prepared by the method described in the previous study. In the wastewater treatment system, nitrification of ammonia component is generally known as rate controlling step. To enhance the rate of nitrification, firstly nitrifying strains Nitrosomonas europaea(IFO14298), are permeabilized chemically, and immobilized on polystyrene films and secondly oxidation rates of strain system and permeabilized strain system are compared in the same condition. with 30 minute permeabilized cells, it took about 25 hours to oxidize 70% of ammonia in the solution, while it took about 40 hours to treat same amount of ammonia with untreated cells. All the immobilization procedures did not harm to the enzyme activity and no mass transfer resistance through the capsule well was shown. In the durability test of immobilized system, the system showed considerable activity for the repeated operation for 90 days. With these results, the system developed in this study showed the possibility to be used in the actual waste water treatment system.

• Journal of Microbiology and Biotechnology
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• 제19권4호
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• pp.346-350
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• 2009

Although salt is known to influence the performance of nitrification significantly, it has not been well reported on how salt affects ammonia-oxidizing bacterial(AOB) community compositions and dynamics in wastewater treatment bioreactors. In this study, these questions were evaluated in a full-scale bioreactor treating saline wastewater. Clone library analysis for the ammonia monooxygenase subunit A gene revealed that AOB belonging to the Nitrosomonas europaea and the N. oligotropha lineages inhabited in the bioreactor. Terminal restriction fragment length polymorphism analysis for monthly samples demonstrated a fluctuation pattern among AOB populations, although AOB within the N. europaea lineage were dominant during the test period. Correlation analysis between patterns of terminal restriction fragments and environmental variables suggested that sodium, chloride, and sulfate were less important; rather, temperature was the most significant factor affecting the AOB community in the bioreactor.

• 미생물학회지
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• 제45권2호
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• pp.112-118
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• 2009

질소제거 하수고도처리공정에서 암모니아산화균은 질소제거에 핵심 역할을 하는 독립영양세균이다. 하수처리 생물반응기에는 다양한 암모니아산화균이 서식하며 군집조성도 시간에 따라 변화한다. 본 연구에서는 생물반응기의 운전인자 및 환경조건이 암모니아산화균 군집구조의 조성과 다양성에 영향을 미친다는 가설을 설정하였다. 이 가설을 검증하기 위해 질산화 반응이 활발한 포항, Palo Alto, Nine Springs, Marshall 하수처리장 활성슬러지 생물반응기로부터 암모니아산화균의 ammonia monooxygenase subunit A 유전자 clone library를 제작하였다. 하수처리 생물반응기에는 Nitrosomonas europaea, N. oligotropha, N.-like, Nitrosospira lineage에 속하는 암모니아산화균이 주로 발견되었으며, N. communis, N. marina, N. cryotolerans lineage에 속하는 암모니아산화균은 주종을 이루지 못했다. 암모니아산화균 군집조성은 하수처리장별로 차이를 보였는데, 포항, Palo Alto, Marshall 하수처리장에서는 N. oligotropha lineage에 속하는 암모니아산화균이 가장 빈번히 발견되었고, Nine Springs 하수처리장에서는 N. europaea lineage에 속하는 암모니아산화균이 주종을 이루었다. 한편, 암모니아산화균 군집조성과 생물반응기 운전인자(HRT, SRT, MLSS) 및 환경조건(온도, pH, COD, $NH_3$, $NO_3{^-}$)의 연관성은 다변수 통계분석법인 Redundancy Analysis 방법을 이용하여 분석하였다. 그 결과, 생물반응조의 COD와 $NO_3{^-}$ 농도가 하수처리 생물반응기에서 암모니아산화균 군집구조를 결정하는 통계학적으로 유의한 변수로 나타났다.

• Biotechnology and Bioprocess Engineering:BBE
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• 제7권5호
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• pp.323-326
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• 2002

To detect whole ammonia-oxidizing bacteria in the activated sludge, group-specific primers targeting the 16S-rRNA gene of ammonia-oxidizing bacteria were used. The electrophoresis pattern of the PCR products seemed to produce a single band of approximately 1.0 k bp for the bacteria in activated sludge and Nitrosomonas europaea. No band was observed for nitrite-oxidizer Nitrobacter winogradskyi and heterotrophs such as Pseudomonas putida. Then direct measurement of the PCR product was made by fluorometry using the reagent Hoechist 33258, so that the fluorescent intensity was in proportional to the cell number of the sample up to 240. Total time required for the test was about 4 h including DNA extraction. The DNA fragments produced were cloned and their sequences showed high similarity to those of Nitrosomonas spp. This study showed the feasibility to detect ammonia-oxidizing bacteria and to esti-mate their population rapidly for the control of the nitrogen elimination process.

• Environmental Engineering Research
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• 제21권2호
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• pp.140-144
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• 2016

Owing to the kinetic differences in ammonia oxidation among ammonia-oxidizing microorganisms (AOM), there is no standard set of kinetic values that can be used as a representative set for nitrifying wastewater treatment plant (WWTP) design. As a result, this study clarified a link between the half-saturation constants for ammonia oxidation (Ks) and the dominant ammonia-oxidizing bacterial (AOB) groups in sludge from full-scale WWTPs and laboratory-scale nitrifying reactors. Quantitative polymerase chain reaction analyses revealed that AOB affiliated with the Nitrosomonas oligotropha cluster were the dominant AOM groups in the sludge taken from the low-ammonia-level WWTPs, while AOB associate with the Nitrosomonas europaea cluster comprised the majority of AOM groups in the sludge taken from the high-ammonia-level WWTPs and nitrifying reactors. A respirometric assay demonstrated that the ammonia Ks values for the high-ammonia-level WWTPs and nitrifying reactors were higher than those of the low-ammonia-level plants. Using the Ks values of available AOM cultures as a reference, the Ks values of the analyzed sludge were mainly influenced by the dominant AOB species. These findings implied that.different sets of kinetic values may be required for WWTPs with different dominant AOM species for more accurate WWTP design and operations.

• Journal of Microbiology and Biotechnology
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• 제17권7호
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• pp.1183-1190
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• 2007

A metabolic uncoupler, 3,3',4',5-tetrachlorosalicylanilide (TCS), was used to reduce excess sludge production in biological wastewater treatment processes. Batch experiments confirmed that 0.4 mg/l of TCS reduced the aerobic growth yield of activated sludge by over 60%. However, the growth yield remained virtually constant even at the increased concentrations of TCS when cultivations were carried out under the anoxic condition. Reduction of sludge production yield was confirmed in a laboratory-scale anoxic-oxic process operated for 6 months. However, it was found that ammonia oxidation efficiency was reduced by as much as 77% in the presence of 0.8 mg/l of TCS in the batch culture. Similar results were also obtained through batch inhibition tests with activated sludges and by bioluminescence assays using a recombinant Nitrosomonas europaea (pMJ217). Because of this inhibitory effect of TCS on nitrification, the TCS-fed continuous system failed to remove ammonia in the influent. When TCS feeding was stopped, the nitrification yield of the process was resumed. Therefore, it seems to be necessary to assess the nitrogen content of wastewater if TCS is used for reducing sludge generation.

• 한국물환경학회지
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• 제22권6호
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• pp.1014-1021
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• 2006

SMMIAR (Submerged Moving Media Intermittent Aeration Reactor) Process is a very efficient system which remove ammonia to nitrogen gas in one reactor. In this study, we determined the diversity of ammonia oxidizing bacteria and denitrifying bacteria using specific PCR amplification and the clone library construction. An ammonia monooxygenase gene(amoA) was analyzed to investigate the diversity of nitrifiers. Most of amoA gene fragments (27/29, 93%) were same types and they are very similar (>99%) to the sequences of Nitrosomonas europaea and other clones isolated from anoxic ammonia oxidizing reactors. ANAMMOX related bacteria have not determined by specific PCR amplification. A nitrite reductase gene(nirK) was analyzed to investigate the diversity of denitrifying bacteria. About half (9/20, 45%) of denitrifiers were clustered with Rhodobacter and most of others were clustered with Mesorhizobium (6/20, 30%) and Rhizobium (3/20, 15%). All of these nirK gene clones were clustered in alpha-Proteobacteria and this result is coincide with other system which also operate nitrification and denitrification in one reactor. The molecular monitoring of the population of nitrifiers and denitrifiers would be helpful for the system stabilization and scale-up.

• Environmental Engineering Research
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• 제23권1호
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• pp.28-35
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• 2018

$17{\alpha}-ethinylestradiol$ (EE2), a synthetic estrogen which interfere the endocrine and reproductive function in living organisms, has been found extensively to be deposited into municipal wastewater treatment plants and the environment via human excretion. EE2 has long been known to be efficiently cometabolized by ammonia-oxidizing bacteria (AOB) during ammonia ($NH_3$) oxidation. Current study aims to investigate the effect of culture history on the biotransformation of EE2 by nitrifying sludge which was enriched under different ammonia loading rates in continuous flow reactors. Result showed that past growth condition largely affected not only the metabolic rate of $NH_3$ oxidation but also EE2 cometabolism. Sludge previously acclimated with higher $NH_3$ loads as well as sludge dominated with AOB belong to high growth cluster (Nitrosomonas europaea-Nitrosococcus mobilis) showed higher rate of EE2 biotransformation than those one being acclimated with lower $NH_3$ loads because of its ability to provide more reducing power from $NH_3$ oxidation. Moreover, the correlation between the degradation rates of $NH_3$ and EE2 was higher in sludge being acclimated with higher load of $NH_3$ in comparison with other sludge. Implication of the findings emphasized the role of volumetric $NH_3$ loading rate in determining EE2 removal in wastewater treatment system.

• 한국미생물·생명공학회지
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• 제35권3호
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• pp.177-183
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• 2007

열충격 시그마인자를 코딩하는 유전자 rpoH가 결여된 돌연변이체 대장균(Escherichia coli satrain A7448)을, 메탄올 자화세균인 Methylovorus sp. strain SS1 DSM11726의 phagemid library로 형질전환 시켜서 $30^{\circ}C$에서 성장하는 Escherichia coli strain A7448 로부터 Methylovorus sp. strain SS1 DSM11726의 rpoH 유전자를 클로닝하고 그 염기서열을 분석하였다. 1,793-bp 염기서열 분석 결과 Methylovorus sp. strain SS1 DSM11726의 RpoH는 284개의 아미노산으로 이루어져 있었으며 예상된 분자량은 32,006, p1값은 5.79로 나타났으며, 동일계열의 ${\beta}$-proteobacteria에 속하는 세균들의 RpoH와 높은 상동성을 보여주었다. Methylovorus sp. strain SS1 DSM11726의 RpoH는 대장균의 RpoH의 기능을 대신할 수 있음을 보여주었다. 열충격 후 RpoH양은 15분까지 지속적으로 증가하다 20분 뒤 양이 감소하는 양상을 나타내었다. 이는 Methylovorus sp. strain SS1 DSM11726의 RpoH 단백질 역시 열에 의해 유도됨을 말해 준다.