• 미생물학회지
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• 제43권3호
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• pp.232-235
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• 2007

해양 및 토양에서의 암모니아 산화는 세균에 비해 Crenarchaeota 그룹의 archaea에 의해 우세하게 일어나고 있음이 알려졌다. 서해 갯벌에서, 배양에 의존하지 알고, 특정 암모니아 산화유전자(amoA)를 가진 archaea을 동정하고자 디지털 PCR법을 응용한 nested PCR법을 개발하였다. amoA와 16S rRNA유전자가 동시에 증폭된 샘플의 분석결과, 16S rRNA유전자에 비해 amoA 유전자의 다양성 이 높았으며, I.1a 그룹의 crenarchaea가 I.1b 그룹의 crenarchaea보다 갯벌지역에서 암모니아 산화에 우점적으로 기여하고 있음을 알 수 있었다. 본 연구에서 시도된, 디지털 PCR과 multiplex-nested PCR을 접목한 접근법을 이용하면 특정 기능유전자를 가진 미생물을 환경에서 검증하는데 응용할 수 있을 것이다.

• 미생물학회지
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• 제50권2호
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• pp.164-168
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• 2014

북극지역은 지구온난화로 인하여 생태계에 큰 영향을 받고 있다. 본 연구는 북극 Svalbard 지역에서 육지 빙하의 해빙(ice melt)의 영향을 받는 해양퇴적층에서 질산화 과정에 핵심역할을 하는 고세균의 질산화유전자(ammonia monooxygenase, AMO)의 공간적 분포의 변화를 조사하였다. 해빙으로 인한 퇴적물 퇴적속도와 고세균 AMO의 alpha subunit를 코딩하는 amoA 유전자와의 관계를 클론라이브러리 분석을 통하여 분석하였다. 육지와 근접하여 퇴적속도가 가장 빠른 정점(188)에서 고세균 amoA 유전자의 다양성이 육지에서 비교적 먼 지역의 정점(176과 184)에 비해 현저히 낮음을 알 수 있었다. 3 정점의 고세균 amoA 유전자 클론들의 평균 아미노산 서열의 상동성은 94%(염기서열 91%)로 나타났다. 176과 184 정점에서 분석된 고세균 amoA 유전자 클론들 중 약 45%가 Nitosopumilus clade와 근연관계에 있는 반면, 188 지역의 경우 낮은 염농도에서 발견되는 Nitrosoarchaeaum clade와 근연관계에 있는 클론들이 발견되었다. 토양 고세균유래 amoA 유전자는 육지에 근접하여 해빙에 의한 영향을 가장 많이 받는 188정점에서만 발견이 되었다. 본 연구를 통하여, 해빙으로 인하여 육지로부터 운반되는 퇴적물의 량이 증가함에 따라, 해양퇴적층의 질소순환관련 미생물 군집에 변화가 유발되는 것으로 추정되며, 고세균의 amoA 유전자가 해양퇴적층 질소순환생태계 변화의 지표로 이용될 수 있음을 알 수 있었다.

• Journal of Microbiology and Biotechnology
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• 제21권6호
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• pp.567-573
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• 2011

We investigated the phylogenetic diversity of ammoniaoxidizing bacteria (AOB) in Yellow Sea continental shelf sediment by the cloning and sequencing of PCR-amplified amoA and 16S rRNA genes. Phylogenetic analysis of the amoA-related clones revealed that the diversity of AOB was extremely low at the study site. The majority (92.7%) of amoA clones obtained belonged to a single cluster, environmental amoA cluster-3, the taxonomic position of which was previously unknown. Phylogenetic analysis on AOB-specific 16S rRNA gene sequences also demonstrated a very low diversity. All of the cloned 16S rRNA gene sequences comprised a single phylotype that belonged to the members of uncultured Nitrosospira cluster-1, suggesting that AOB belonging to the uncultured Nitrosospira cluster-1 could carry amoA sequences of environmental amoA cluster-3.

• 한국미생물·생명공학회지
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• 제47권2호
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• pp.296-302
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• 2019

In this study, the effects of ammonium concentration ($117.5-1155.0mg-N{\cdot}l^{-1}$), nitrite concentration ($0-50.0mg-N{\cdot}l^{-1}$), and temperature ($15-35^{\circ}C$) on nitrification performance and its functional genes (amoA-arc, amoA-bac, hao) in an enriched consortium inoculated with humic acid were determined. Notably, the maximum nitrification rate value was observed at $315mg-N{\cdot}l^{-1}$ of ammonium, but the highest functional gene copy numbers were obtained at $630mg-N{\cdot}l^{-1}$ of ammonium. No inhibition of the nitrification rate and functional gene copy numbers was observed via the added nitrites. The optimum temperature for maximum nitrification performance was observed to be $30^{\circ}C$. The amoA-bac copy numbers were also greater than those of amoA-arc under all test conditions. Notably, amoA-arc copy numbers and nitrification efficiency showed a positive relationship in network analysis. These results indicate that ammonium-oxidizing archaea and bacteria play important roles in the nitrification process.

• 미생물학회지
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• 제51권4호
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• pp.329-337
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• 2015

Nitrosomonadales 목에서 속하는 균주 중 현재 유전체 서열이 알려진 모든 유전체(N=10)를 이용하여 범유전체 및 핵심유전체 분석을 수행한 결과, 각각 9,808개와 908개 유전자클러스터를 포함하는 것을 확인하였다. Betaproteobacteria의 다른 목의 참조군들과 비교를 통하여 범유전체와 핵심유전체의 크기에 유전체의 수와 집단 내의 유전체들의 차이가 영향을 미치는 것을 확인하였다. Nitrosomonas 속과 Nitrosospira 속의 범유전체는 7,180개와 4,586개, 핵심유전체는 1,092개와 1,600로로 각각 측정되어 Nitrosospira 속의 동질성이 더 높은 것을 확인하였다. Nitrosomonadales 목의 범유전체와 핵심유전체의 크기에 Nitrosomonas 속이 대부분의 영향을 미치는 것을 확인하였다. COG 분석을 통하여 핵심유전체의 크기에는 J (translation, ribosomal structure and biogenesis) 범주가 가장 큰 비율(9.7-21.0%)을 차지하며, 유전체 사이의 유전적 거리가 먼 집단일수록 그 비율이 높아지는 것을 확인하였다. 범유전체의 크기에는 "-" (unclassified) 범주가 34-51%의 높은 비율을 차지하고 있을 정도로 큰 영향을 미치는 것을 확인하였다. 총 97개의 유전자 클러스터가 참조군에는 없고 Nitrosomonadales에만 존재하는 것을 확인하였다. 이들 클러스터들은 Nitrosomonadales을 특징 지우는 유전자들인 ammonia monooxygenase의 유전자인 amoA와 amoB와 그와 관련 있는 amoE와 amoD들을 포함하는 반면에 unclassified 유전자들도 상당량(16-45%)을 포함하고 있다. 이러한 유전자 클러스터는 Nitrosomonadales의 유전적 특이성을 밝히는 데 중요한 역할을 할 것이다.

• Journal of Microbiology and Biotechnology
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• 제20권7호
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• pp.1128-1133
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• 2010

The diversity and abundance of ammonia-oxidizing bacteria (AOB) in activated sludge were compared using PCR-DGGE and real-time PCR assays. Activated sludge samples were collected from five different types of wastewater treatment plants (WWTPs) mainly treating textile, paper, food, and livestock wastewater or domestic sewage. The composition of total bacteria determined by PCR-DGGE was highly diverse between the samples, whereas the community of AOB was similar across all the investigated activated sludge. Total bacterial numbers and AOB numbers in the aerated mixed liquor were in the range of $1.8{\times}10^{10}$ to $3.8{\times}10^{12}$ and $1.7{\times}10^6$ to $2.7{\times}10^{10}$ copies/l, respectively. Activated sludge from livestock, textile, and sewage treating WWTPs contained relatively high amoA gene copies (more than $10^5$ copies/l), whereas activated sludge from food and paper WWTPs revealed a low number of the amoA gene (less than $10^3$ copies/l). The value of the amoA gene copy effectively showed the difference in composition of bacteria in different activated sludge samples and this was better than the measurement with the AOB 16S rRNA or total 16S rRNA gene. These results suggest that the quantification of the amoA gene can help monitor AOB and ammonia oxidation in WWTPs.

• Environmental Engineering Research
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• 제20권1호
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• pp.105-109
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• 2015

Triclosan is a synthetic antimicrobial agent used in numerous industrial and personal care products. Triclosan collected in wastewater treatment plants can be biodegraded up to 80%. However, little is studied about the abundances of known triclosan-degrading bacteria in activated sludge systems. A previous study reported that Sphingopyxis strain KCY1 isolated from activate sludge can cometabolically degrade triclosan. Recently, a quantitative PCR (qPCR) assay specific to strain KCY1 has been developed. Thus, this study investigated the abundance of strain KCY1 in three different activated sludge wastewater treatments using a qPCR assay. Additionally, ammonia-oxidizing bacteria (AOB), known as triclosan-degraders, and amoA gene were quantified. Strain KCY1 were detected in activated sludge samples from three different wastewater treatment plants. The concentrations of strain KCY1 and AOB were on the order of $10^5-10^6$ gene copies/mL, while amoA gene concentration was on the order of $10^4$ gene copies/mL.

• Journal of Microbiology and Biotechnology
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• 제23권9호
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• pp.1187-1196
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• 2013

Global warming will have far-reaching effects on our ecosystem. However, its effects on Antarctic soils have been poorly explored. To assess the effects of warming on microbial abundance and community composition, we sampled Antarctic soils from the King George Island in the Antarctic Peninsula and incubated these soils at elevated temperatures of $5^{\circ}C$ and $8^{\circ}C$ for 14 days. The reduction in total organic carbon and increase in soil respiration were attributed to the increased proliferation of Bacteria, Fungi, and Archaea. Interestingly, bacterial ammonia monooxygenase (amoA) genes were predominant over archaeal amoA, unlike in many other environments reported previously. Phylogenetic analyses of bacterial and archaeal amoA communities via clone libraries revealed that the diversity of amoA genes in Antarctic ammonia-oxidizing prokaryotic communities were temperature-insensitive. Interestingly, our data also showed that the amoA of Antarctic ammonia-oxidizing bacteria (AOB) communities differed from previously described amoA sequences of cultured isolates and clone library sequences, suggesting the presence of novel Antarctic-specific AOB communities. Denitrification-related genes were significantly reduced under warming conditions, whereas the abundance of amoA and nifH increased. Barcoded pyrosequencing of the bacterial 16S rRNA gene revealed that Proteobacteria, Acidobacteria, and Actinobacteria were the major phyla in Antarctic soils and the effect of short-term warming on the bacterial community was not apparent.

• 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.

• 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.