研究者データベース

研究者情報

マスター

アカウント(マスター)

  • 氏名

    岡部 聡(オカベ サトシ), オカベ サトシ

所属(マスター)

  • 工学研究院 環境工学部門 環境工学

所属(マスター)

  • 工学研究院 環境工学部門 環境工学

独自項目

syllabus

  • 2020, 大学院共通授業科目(教育プログラム):JICA開発大学院連携プログラム, Inter-Graduate School Classes(Educational Program):JICA Development Study Program, 修士課程, 大学院共通科目, Japan's experience for resources development and environmental conservation, Japan's ODA, Technical cooperation project, Grant aid project, Yen loan project
  • 2020, 環境微生物工学特論, Environmental Biotechnology, 修士課程, 工学院, microbiology, molecular biology, microbial community analysis, microbial ecology, water and wastewater treatment, biofilm process
  • 2020, 環境微生物工学特論, Environmental Biotechnology, 博士後期課程, 工学院, microbiology, molecular biology, microbial community analysis, microbial ecology, water and wastewater treatment, biofilm process
  • 2020, 一般教育演習(フレッシュマンセミナー), Freshman Seminar, 学士課程, 全学教育, 水問題、都市水循環、水環境保全、水処理技術、水の再利用、水の安全性評価、水のマネージメント
  • 2020, 上水工学, Water Works Engineering, 学士課程, 工学部, 水資源、水循環、上水道計画、浄水技術、高度浄水処理
  • 2020, 微生物工学, Environmental Microbiology, 学士課程, 工学部, バクテリア、微生物生態、微生物反応、反応動力学、遺伝子工学、環境浄化 

timetable

  • 修士課程, 工学院, 2020, 水・公衆衛生特論
  • 修士課程, 工学院, 2020, Environmental Biotechnology(環境バイオテクノロジーE)
  • 博士後期課程, 工学院, 2020, 水・公衆衛生特論
  • 博士後期課程, 工学院, 2020, Environmental Biotechnology(環境バイオテクノロジーE)

researchmap

プロフィール情報

学位

  • Ph.D(米国Montana州立大学)

プロフィール情報

  • 岡部, オカベ
  • 聡, サトシ
  • ID各種

    200901068708757596

対象リソース

業績リスト

研究キーワード

  • 微生物燃料電池(MFC)   嫌気性アンモニア酸化(anammox)   分子生物学   環境微生物生態学   糞便汚染   病原微生物   水質保全   水の安全性評価   浄水処理   廃水処理   Molecular microbiology   Environmental Microbiology   fecal pollution   pathogenic bacteria   Water pollution contorl   Water safety assessment   Water Treatment   Wastewater Treatment   

研究分野

  • ライフサイエンス / 生態学、環境学
  • 環境・農学 / 環境材料、リサイクル技術
  • 環境・農学 / 環境負荷低減技術、保全修復技術
  • 社会基盤(土木・建築・防災) / 土木環境システム

経歴

  • 2008年04月 - 現在 北海道大学大学院 工学研究院環境創生工学部門 教授
  • 2000年04月 - 2008年03月 北海道大学大学院 都市環境工学専攻 助教授
  • 1994年04月 - 2000年03月 北海道大学 工学部衛生工学科 助手
  • 1993年01月 - 1994年03月 宮崎大学 工学部土木環境工学科 助手

学歴

  •         - 1992年12月   米国Montana州立大学   大学院博士課程修了   土木工学専攻

委員歴

  • 2021年04月 - 現在   土木学会環境工学委員会   委員長
  • 2021年01月 - 現在   日本微生物生態学会   会長
  • 2020年10月 - 現在   日本学術会議   連携会員
  • 2019年06月 - 現在   日本水環境学会   理事
  • 2017年02月 - 現在   International Society for Microbial Ecology   Senior Editor
  • 2010年01月 - 現在   Water Science and Technology   Associate Editor
  • 2001年 - 現在   「Environmental Technology」編集委員   編集委員   「Environmental Technology」編集委員
  • 2001年 - 現在   「Biodegradation」編集委員   編集委員   「Biodegradation」編集委員
  • 2012年08月 - 2017年08月   International Society for Microbial Ecology (ISME)   International Borad member
  • 2004年   International Water Association   Biofilm specialist group management committee   International Water Association
  • 2003年   International Water Association   Activated Sludge Ppulation Dynamics management committee   International Water Association

受賞

  • 2018年03月 北海道大学 北海道大学教育研究総長表彰(研究部門)
     
    受賞者: 岡部 聡
  • 2017年03月 北海道大学 北海道大学教育研究総長表彰(教育部門)
     
    受賞者: 岡部 聡
  • 2016年 International Water Association IWA Fellow
     
    受賞者: 岡部 聡
  • 2014年04月 Nagase Science and Technology Foundation 長瀬研究振興賞
     
    受賞者: 岡部 聡
  • 2009年 日本水環境学会論文賞
  • 2008年 日本学術振興会賞
  • 2008年 JSPS prize
  • 2006年 The best poster award at The 6th International Conference on Biofilm Systems
  • 2005年 土木学会環境工学フォーラム優秀ポスター発表賞
  • 2003年 環境工学フォーラム論文賞
  • 2001年 月刊「水」論文賞
  • 1999年 日本水道協会論文有効賞
  • 1999年 日本水環境学会論文奨励賞(廣瀬賞)
  • 1999年 平成10年度土木学会年次学術講演会優秀講演者賞
  • 1998年 平成9年度土木学会年次学術講演会優秀講演者賞

論文

  • Hiroe Hara-Yamamura, Koji Nakashima, Toshikazu Fukushima, Satoshi Okabe
    Chemosphere 286 Pt 2 131715 - 131715 2021年08月05日 
    The biological impacts of residual pharmaceuticals in the complex wastewater effluents have not been fully understood. Here, we investigated changes in the transcriptomic responses of hepatobrastoma (HepG2) cells exposed to a single or partially combined three common non-steroidal anti-inflammatory drugs (NSAIDs); ketoprofen (KPF), mefenamic acid (MFA) and diclofenac (DCF), in domestic wastewater effluents. After 48 h sub-lethal exposure to single compounds, the DNA microarray analysis identified 57-184 differently expressed genes (DEGs). The hierarchical clustering analysis and GO enrichment of the DEGs showed that gene expression profiles of the NSAIDs were distinct from each other although they are classified into the same therapeutic category. Four maker genes (i.e., EGR1, AQP3, SQSTM1, and NAG1) were further selected from the common DEGs, and their expressions were quantified by qPCR assay in a dose-dependent manner (ranging from μg/L to mg/L). The results revealed the insignificant induction of the marker genes at 1 μg/L of KPF, MFA, and DCF, suggesting negligible biological impacts of the NSAIDs on gene expression (early cellular responses) of HepG2 at typical concentration levels found in the actual wastewater effluents. Based on the quantitative expression analysis of the selected marker genes, the present study indicated that the presence of wastewater effluent matrix may mitigate the potentially adverse cellular impacts of the NSAIDs.
  • N'Dah Joel Koffi, Satoshi Okabe
    Chemosphere 274 129715 - 129715 2021年07月 
    Nitrogen removal from wastewater is an indispensable but highly energy-demanding process, and thus more energy-saving treatment processes are required. Here, we investigated the performance of bioelectrochemical ammonium nitrogen (NH4+-N) removal from real domestic wastewater without energy-intensive aeration by a single chamber microbial electrolysis cell (MEC) that was electrically powered by a double chamber microbial fuel cell (MFC). Anoxic NH4+-N oxidation and total nitrogen (TN) removal rates were determined at various applied voltages (0-1.2 V), provided by the MFC. The MEC achieved a NH4+-N oxidation rate of 151 ± 42 g NH4+-N m-3 d-1 and TN removal rate of 95 ± 42 g-TN m-3 d-1 without aeration at the applied voltage of 0.8 V (the anode potential Eanode = +0.633 ± 0.218 V vs. SHE). These removal rates were much higher than the previously reported values and conventional biological nitrogen removal processes. Open and closed-circuit MEC batch experiments confirmed that anoxic NH4+-N oxidation was an electrochemically mediated biological process (that is, an anode acted as an electron acceptor) and denitrification occurred simultaneously without NO2- and NO3- accumulation. Moreover, ex-situ15N tracer experiment and microbial community analysis revealed that anammox and heterotrophic denitrification mainly contributed to the TN removal. Thus, the bioelectrochemical anodic NH4+-N oxidation was coupled with anammox and denitrification in this MFC-assisted MEC system. Taken together, our MFC-driven single chamber MEC could be a high rate energy-saving nitrogen removal process without external carbon and energy input and high energy-demanding aeration.
  • Satoshi Okabe, Atsushi Kamigaito, Kanae Kobayashi
    The ISME journal 2021年06月18日 
    Little is known about the cell physiology of anammox bacteria growing at extremely low growth rates. Here, "Candidatus Brocadia sinica" and "Candidatus Scalindua sp." were grown in continuous anaerobic membrane bioreactors (MBRs) with complete biomass retention to determine maintenance energy (i.e., power) requirements at near-zero growth rates. After prolonged retentostat cultivations, the specific growth rates (μ) of "Ca. B. sinica" and "Ca. Scalindua sp." decreased to 0.000023 h-1 (doubling time of 1255 days) and 0.000157 h-1 (184 days), respectively. Under these near-zero growth conditions, substrate was continuously utilized to meet maintenance energy demands (me) of 6.7 ± 0.7 and 4.3 ± 0.7 kJ mole of biomass-C-1 h-1 for "Ca. B. sinica" and "Ca. Scalindua sp.", which accorded with the theoretically predicted values of all anaerobic microorganisms (9.7 and 4.4 kJ mole of biomass-C-1 h-1at 37 °C and 28 °C, respectively). These me values correspond to 13.4 × 10-15 and 8.6 × 10-15 watts cell-1 for "Ca. B. sinica" and "Ca. Scalindua sp.", which were five orders of magnitude higher than the basal power limit for natural settings (1.9 × 10-19 watts cells-1). Furthermore, the minimum substrate concentrations required for growth (Smin) were calculated to be 3.69 ± 0.21 and 0.09 ± 0.05 μM NO2- for "Ca. B. sinica" and "Ca. Scalindua sp.", respectively. These results match the evidence that "Ca. Scalindua sp." with lower maintenance power requirement and Smin are better adapted to energy-limited natural environments than "Ca. B. sinica", suggesting the importance of these parameters on ecological niche differentiation in natural environments.
  • Satoshi Okabe, Amrini Amalia Shafdar, Kanae Kobayashi, Lei Zhang, Mamoru Oshiki
    The ISME journal 15 5 1287 - 1301 2021年05月 
    Presence of glycogen granules in anaerobic ammonium-oxidizing (anammox) bacteria has been reported so far. However, very little is known about their glycogen metabolism and the exact roles. Here, we studied the glycogen metabolism in "Ca. Brocadia sinica" growing in continuous retentostat cultures with bicarbonate as a carbon source. The effect of the culture growth phase was investigated. During the growing phase, intracellular glycogen content increased up to 32.6 mg-glucose (g-biomass dry wt)-1 while the specific growth rate and ATP/ADP ratio decreased. The accumulated glycogen begun to decrease at the onset of entering the near-zero growth phase and was consumed rapidly when substrates were depleted. This clearly indicates that glycogen was synthesized and utilized as an energy storage. The proteomic analysis revealed that "Ca. B. sinica" synthesized glycogen via three known glycogen biosynthesis pathways and simultaneously degraded during the progress of active anammox, implying that glycogen is being continuously recycled. When cells were starved, a part of stored glycogen was converted to trehalose, a potential stress protectant. This suggests that glycogen serves at least as a primary carbon source of trehalose synthesis for survival. This study provides the first physiological evidence of glycogen metabolism in anammox bacteria and its significance in survival under natural substrate-limited habitat.
  • Daisuke Sano, Ryosuke Watanabe, Wakana Oishi, Mohan Amarasiri, Masaaki Kitajima, Satoshi Okabe
    Food and environmental virology 13 1 84 - 92 2021年03月 
    This study investigated the influence of viral interference on the detection of enteric viruses using the integrated cell culture (ICC)-PCR with a BGM cell line. It was possible to detect 102 plaque-forming units (PFU)/flask of enterovirus 71 (EV71) in spite of the presence of 104 PFU/flask of adenovirus 40 (AdV40). Meanwhile, 104 PFU/flask of AdV40 was not detected in the presence of 102 PFU/flask of EV71. This inhibition of AdV40 detection using ICC-PCR was attributable to the growth of EV71, because the addition of a growth inhibitor of EV71 (rupintrivir) neutralized the detection inhibition of AdV40. The growth inhibition of AdV40 under co-infection with EV71 is probably caused by the immune responses of EV71-infected cells. AdV is frequently used as a fecal contamination indicator of environmental water, but this study demonstrated that false-negative detection of infectious AdV using ICC-PCR could be caused by the co-existence of infectious EV in a water sample. The addition of rupintrivir could prevent false-negative detection of AdV using ICC-PCR. This study, therefore, emphasizes the importance of confirming the presence of multiple enteric viruses in a sample derived from environmental water prior to the application of ICC-PCR because the viral interference phenomenon may lead to the false-negative detection of target viruses.
  • Meri Nakajima, Reiko Hirano, Satoshi Okabe, Hisashi Satoh
    Chemosphere 263 128331 - 128331 2021年01月 
    Domestic and industrial wastewater treatment systems are vital in the protection of natural ecosystems and human health. Identification of microbial communities in the systems is essential to stable treatment performance. However, the current tools of microbial community analysis are labor intensive and time consuming, and require expensive equipment. Therefore, we developed a simple assay for colorimetric quantification of bacterial 16S rRNA extracted from environmental samples. The assay is based on RNA extraction with commercial kits, mixing the unamplified RNA sample with Au-nanoprobes and NaCl, and analyzing the absorbance spectra. Our experimental results confirmed that the assay format was valid. By analyzing the synthesized DNA, we optimized the operational parameters affecting the assay. We achieved adequate capture DNA density by setting the capture DNA probe concentration at 10 μM during the functionalization step. The required incubation time after NaCl addition was 30 min. The binding site of the target had negligible effect on DNA detection. Under the optimized condition, a calibration curve was created using 16S rRNA extracted from activated sludge. The curve was linear above 5.0 × 107 copies/μL of bacterial 16S rRNA concentration, and the limit of detection was 1.17 × 108 copies/μL. Using the calibration curve, the bacterial 16S rRNA concentration in activated sludge samples could be quantified with deviations between 48% and 208% against those determined by RT-qPCR. The findings of our study introduce an innovative tool for the quantification of 16S rRNA concentration as the activity of key bacteria in wastewater treatment processes, achieving stable treatment performance.
  • N'Dah Joel Koffi, Satoshi Okabe
    Scientific reports 10 1 18985 - 18985 2020年11月04日 
    Although microbial fuel cells (MFCs) can produce renewable energy from wastewater, the generated power is practically unusable. To extract usable power from an MFC fed with wastewater, we newly developed a low voltage booster multiplier (LVBM), which is composed of a self-oscillating LVB and multistage voltage multiplier circuits (VMCs). The low output MFC voltage (ca. 0.4 V) was successfully boosted up to 99 ± 2 V, which was the highest voltage that has been ever reported, without voltage reversal by connecting an LVB with 20-stage VMCs. Moreover, the boosted voltage (81 ± 1 V) was stably maintained for > 40 h even after disconnecting the LVBM from the MFC. The energy harvesting efficiency of LVBM was > 80% when an LVB with 4-stage VMCs was charged to 9.3 V. These results clearly suggest that the proposed LVBM system is an efficient and self-starting energy harvester and storage for low-power generating MFCs.
  • Kanae Kobayashi, Keitaro Fukushima, Yuji Onishi, Kazuya Nishina, Akiko Makabe, Midori Yano, Scott D Wankel, Keisuke Koba, Satoshi Okabe
    Rapid communications in mass spectrometry : RCM 2020年10月14日 [査読有り][通常論文]
     
    RATIONALE: Oxygen isotope ratio measurements of NO2- and NO3- by the azide method and denitrifier method are sensitive to the δ18 O of sample water. However, the influence of δ18 OH2O on those measurements has not been quantitatively evaluated and documented so far. Therefore, we investigated the influence of δ18 OH2O of sample on the δ18 O analysis of NO2- and NO3- . METHODS: We prepared NO2- and NO3- standards (with known δ18 ONO2- and δ18 ONO3- values) dissolved in waters having different δ18 OH2O values (δ18 OH2O = -12.6, 25.9, 56.7, and 110.1‰). Nitrite and nitrate were converted to N2 O using the azide method and the denitrifier method, respectively. The isotope ratios of the generated N2 O were measured with a Sercon PT-GC/IRMS system. The measured δ18 O values of the produced N2 O were plotted against known δ18 ONO2- and δ18 ONO3- values to evaluate the influence of exchange of an oxygen atom with H2 O during the conversion of NO2- to N2 O and NO3- to N2 O, respectively. RESULTS: The degree of O isotope exchange was 10.8 ± 0.3% in the azide method and 5.5 ± 1.0 % in the denitrifier method, indicating that the azide method is more susceptible to artifacts arising from differences in the δ18 OH2O value of water than the denitrifier method. Thus, the intercept of the standard calibration curve must be corrected to account for differences in δ18 OH2O . Abiotic NO2 - H2 O equilibrium isotope effect experiments yielded a rate constant of (1.13 ± 007) × 10-2 (h-1 ) and an equilibrium isotope effect of 11.9 ± 0.1‰ under the condition of pH=7.5, 30°C, and 2.5% salinity. CONCLUSIONS: Oxygen isotope ratio measurements of NO2- by the azide method are highly sensitive to δ18 OH2O as a result of significant oxygen isotope exchange between NO2- and H2 O. Therefore, to obtain the most accurate measurements the same δ18 OH2O value as that of the sample must be used to make the NO2- and NO3- standards.
  • Yifan Zhu, Hiroki Kawai, Satoshi Hashiba, Mohan Amarasiri, Masaaki Kitajima, Satoshi Okabe, Daisuke Sano
    Molecules (Basel, Switzerland) 25 18 2020年09月07日 [査読有り][通常論文]
     
    In this study, we investigated the impact of GD1a-expressing bacterial strains on the infectivity of murine norovirus (MNV). Eligible bacterial strains were screened from a sewage sample using flow cytometry, and their genetic sequences of 16S rRNA were determined. The enzyme-linked immunosorbent assay (ELISA) was employed to analyze the binding between bacteria and MNV particles, and the plaque assay was used to assess the effects of GD1a-positive and negative strains on MNV infectivity. The result from ELISA shows that MNV particles are able to bind to both GD1a-positive and negative bacterial strains, but the binding to the GD1a-positive strain is more significant. The infectivity assay result further shows that the MNV infectious titer declined with an increasing concentration of GD1a-positive bacteria. The addition of anti-GD1a antibody in the infectivity assay led to the recovery of the MNV infectious titer, further confirming that the binding between MNV particles and bacterial GD1a ganglioside compromises MNV infectivity. Our findings highlight the role indigenous bacteria may play in the lifecycle of waterborne enteric viruses as well as the potential of exploiting them for virus transmission intervention and water safety improvement.
  • Kazuki Jokai, Tomomi Nakamura, Satoshi Okabe, Satoshi Ishii
    Chemosphere 262 127838 - 127838 2020年08月01日 [査読有り][通常論文]
     
    Nitrogen and heavy metals can co-occur in various industrial wastewaters such as coke-oven wastewater. Removal of these contaminants is important, but cost-efficient removal technology is limited. In this study, we examined the usefulness of nitrate-dependent ferrous iron oxidation (NDFO) for the simultaneous removal of nitrate and heavy metals (iron and zinc), by using an NDFO strain Pseudogulbenkiania sp. NH8B. Based on the batch culture assays, nitrate, Fe, and Zn were successfully removed from a basal medium as well as coke-oven wastewater containing 5 mM nitrate, 10 mM Fe(II), and 10 mg/L Zn. Zinc in the water was most likely co-precipitated with Fe(III) oxides produced during the NDFO reaction. Simultaneous removal of nitrate, Fe, and Zn was also achieved in a continuous-flow reactor fed with a basal medium containing 10 mM nitrate, 5 mM Fe(II), 4 mM acetate, and 10 mg/L Zn. However, when the reactor is fed with coke-oven wastewater supplemented with 10 mM nitrate, 5 mM Fe(II), 4 mM acetate, and 10 mg/L ZnCl2, the reactor performance significantly decreased, most likely due to the inhibition of bacterial growth by thiocyanate or organic contaminants present in the coke-oven wastewater. Use of mixed culture of NDFO bacteria and thiocyanate/organic-degrading denitrifiers should help improve the reactor performance.
  • Syun-Suke Kadoya, Syun-Ichi Urayama, Takuro Nunoura, Miho Hirai, Yoshihiro Takaki, Masaaki Kitajima, Toyoko Nakagomi, Osamu Nakagomi, Satoshi Okabe, Osamu Nishimura, Daisuke Sano
    Journal of virology 94 10 2020年05月04日 [査読有り][通常論文]
     
    RNA viruses form a dynamic distribution of mutant swarms (termed "quasispecies") due to the accumulation of mutations in the viral genome. The genetic diversity of a viral population is affected by several factors, including a bottleneck effect. Human-to-human transmission exemplifies a bottleneck effect, in that only part of a viral population can reach the next susceptible hosts. In the present study, two lineages of the rhesus rotavirus (RRV) strain of rotavirus A were serially passaged five times at a multiplicity of infection (MOI) of 0.1 or 0.001, and three phenotypes (infectious titer, cell binding ability, and specific growth rate) were used to evaluate the impact of a bottleneck effect on the RRV population. The specific growth rate values of lineages passaged under the stronger bottleneck (MOI of 0.001) were higher after five passages. The nucleotide diversity also increased, which indicated that the mutant swarms of the lineages under the stronger bottleneck effect were expanded through the serial passages. The random distribution of synonymous and nonsynonymous substitutions on rotavirus genome segments indicated that almost all mutations were selectively neutral. Simple simulations revealed that the presence of minor mutants could influence the specific growth rate of a population in a mutant frequency-dependent manner. These results indicate a stronger bottleneck effect can create more sequence spaces for minor sequences.IMPORTANCE In this study, we investigated a bottleneck effect on an RRV population that may drastically affect the viral population structure. RRV populations were serially passaged under two levels of a bottleneck effect, which exemplified human-to-human transmission. As a result, the genetic diversity and specific growth rate of RRV populations increased under the stronger bottleneck effect, which implied that a bottleneck created a new space in a population for minor mutants originally existing in a hidden layer, which includes minor mutations that cannot be distinguished from a sequencing error. The results of this study suggest that the genetic drift caused by a bottleneck in human-to-human transmission explains the random appearance of new genetic lineages causing viral outbreaks, which can be expected according to molecular epidemiology using next-generation sequencing in which the viral genetic diversity within a viral population is investigated.
  • Hisashi Satoh, Kai Kikuchi, Yutaka Katayose, Shu Tsuda, Reiko Hirano, Yuga Hirakata, Masaaki Kitajima, Satoshi Ishii, Mamoru Oshiki, Masashi Hatamoto, Masahiro Takahashi, Satoshi Okabe
    The Science of the total environment 715 136928 - 136928 2020年05月01日 [査読有り][通常論文]
     
    Monitoring of Escherichia coli concentrations at wastewater treatment plants (WWTPs) is important to ensure process performance and protect public health. However, conventional E. coli enumeration methods are complicated and time- and labor-consuming. Here, we report a novel simple and reliable method based on β-d-glucuronidase (GUS) activity assay to enumerate E. coli concentrations in wastewater (WW) samples. An aliquot (20 μL) of the medium with fluorogenic enzyme substrate for E. coli and 180 μL of a WW sample were added to one well of a 96-well microplate. The microplate was placed in a microplate reader at 37 °C. To this end, the fluorescence intensity of a fluorogenic enzyme substrate for E. coli was measured every 10 min over 3 h to determine GUS activity. The linear increase in the fluorescence intensity representing the GUS activities showed a positive correlation with E. coli concentrations in wastewater samples. However, the correlation equations were specific to WWTPs, which could be due to the difference in the E. coli population structures among WWTPs. We observed that the wastewater matrix is not a limitation to measure the GUS activity, and a WWTP-specific correlation equation can be used as a calibration curve to estimate the E. coli concentrations in the samples collected from that site. A comparison of the results with those of culture-dependent Colilert method proved that the current method is simple and useful for the enumeration of E. coli concentrations in wastewater samples reliably.
  • Lei Zhang, Satoshi Okabe
    Water research 171 115468 - 115468 2020年03月15日 [査読有り][通常論文]
     
    Anaerobic ammonium oxidizing (anammox) bacteria can directly convert ammonium and nitrite to nitrogen gas anaerobically and were responsible for a substantial part of the fixed nitrogen loss and re-oxidation of nitrite to nitrate in freshwater and marine ecosystems. Although a wide variety of studies have been undertaken to investigate the abundance and biodiversity of anammox bacteria so far, ecological niche differentiation of anammox bacteria is still not fully understood. To assess their growth behavior and consequent population dynamics at a given environment, the Monod model is often used. Here, we summarize the Monod kinetic parameters such as the maximum specific growth rate (μmax) and the half-saturation constant for nitrite (KNO2-) and ammonium (KNH4+) of five known candidatus genera of anammox bacteria. We also discuss potential pivotal environmental factors and metabolic flexibility that influence the community compositions of anammox bacteria. Particularly biodiversity of the genus "Scalindua" might have been largely underestimated. Several anammox bacteria have been successfully enriched from various source of biomass. We reevaluate their enrichment methods and culture medium compositions to gain a clue of niche differentiation of anammox bacteria. Furthermore, we formulate the current issues that must be addressed. Overall this review re-emphasizes the importance of enrichment cultures (preferably pure cultures), physiological characterization and direct microbial competition studies using enrichment cultures in laboratories.
  • Andri Taruna Rachmadi, Masaaki Kitajima, Tsuyoshi Kato, Hiroyuki Kato, Satoshi Okabe, Daisuke Sano
    Environmental science & technology 54 4 2068 - 2077 2020年02月18日 [査読有り][通常論文]
     
    A credit value of virus inactivation has been assigned to the disinfection step in international and domestic guidelines for wastewater reclamation and reuse. To fulfill the credit value for water disinfection, water engineers need to apply an appropriate disinfection strength, expressed as a CT value (mg × min/L), which is a product of disinfectant concentration and contact time, against enteric viruses in wastewater. In the present study, we extracted published experimental data on enteric virus inactivation using free chlorine and monochloramine and applied the Tobit analysis and simple linear regression analysis to calculate the range of CT values (mg × min/L) needed for 4-log10 inactivation. Data were selected from peer-reviewed papers containing kinetics data of virus infectivity and chlorine residual in water. Coxsackie B virus and echovirus require higher CT values (lower susceptibility) for 4-log10 inactivation than adenovirus and a human norovirus surrogate (murine norovirus) with free chlorine. On the other hand, adenovirus has lower susceptibility to monochloramine compared to murine norovirus, coxsackievirus, and echovirus. The factors that influence the required CT value are virus type, pH, water temperature, and water matrix. This systematic review demonstrates that enteroviruses and adenovirus are appropriate representative enteric viruses to evaluate water disinfection using free chlorine and monochloramine, respectively.
  • Hiroe Hara-Yamamura, Toshikazu Fukushima, Lea Chua Tan, Satoshi Okabe
    Chemosphere 240 124894 - 124894 2020年02月 [査読有り][通常論文]
     
    We performed a transcriptome-based bioassay (TSB assay) using human hepatoma HepG2 cells to evaluate the potential toxicity of whole wastewater effluents from two membrane bioreactors (MBRs) and a conventional activated sludge process (AS). The biologically active agent(s) in the wastewater effluents were characterized based on expression of the marker genes (i.e., CYP1A1, AKR1B10, GCLM and GPX2) selected by DNA microarray analysis, after the wastewater effluent samples were concentrated by a reverse osmosis (RO) membrane and further fractionated by various manipulations. The qPCR assay of marker genes demonstrated that the induction of CYP1A1 and GPX2 was mitigated after passing through C18 and chelate columns. In addition, clear induction of CYP1A1 was observed in the smallest size fraction with 1 k Da or smaller organic molecules in all the tested effluents. These results together with the water quality data of the fractionated samples suggested that responsible constituents for potentially adverse and abnormal transcriptomic responses in HepG2 could have hydrophobic nature and act with metal-dissolved organic matter (DOM) complexes in 1 k Da or smaller size fraction. Although DOM is known to play two contradictory roles as a protector and an inducer of toxicants, our present study indicated the DOM in wastewater effluent, particularly humic substances with acidic nature, functioned as a toxicity inducer of residual chemicals in the effluents. This study provided a new insight into the nature of "toxic unknowns" in the wastewater effluents, which should be monitored whole through the reclamation process and prioritized for removal.
  • Toshikazu Fukushima, Wongta Jintana, Satoshi Okabe
    Environmental science and pollution research international 27 6 6326 - 6337 2020年02月 [査読有り][通常論文]
     
    Although toxicity of silver nanoparticles (AgNPs) has been well studied, the mixture toxicity of the combination of AgNPs and other environmental pollutants is still largely unknown. Here, we investigated the mixture toxicity of the combinations of AgNPs and common environmental pollutants such as arsenic (As), cadmium (Cd), and chromium (Cr) on human hepatoma cell line (HepG2) at noncytotoxic concentrations based on analyses of cytotoxicity, genotoxicity, reactive oxygen species (ROS) generation, and modes of cell death. In addition, DNA microarray analysis was performed to understand the cellular responses at a molecular level. AgNPs-As and AgNPs-Cd combinations exhibited synergistic effect on cytotoxicity while AgNPs-Cr showed additive effect. The AgNPs-Cd combination caused much stronger synergism than AgNPs-As combination. Based on cellular and molecular level analyses, the synergistic effect could be explained by overproduction of reactive oxygen species (ROS), which induced DNA damage and consequently apoptotic cell death. On the other hand, the additive effect caused by AgNPs-Cr could be attributed to reduction of the mixture toxicity by precipitation of Cr ions. Taken together, our results clearly demonstrated that the mixture toxicity of AgNPs with As, Cd, or Cr at noncytotoxic concentrations had different toxicity effects. Particularly, toxicogenomic approach using DNA microarray was useful to assess the mechanisms of the mixture toxicity.
  • Mamoru Oshiki, Haruna Hiraizumi, Hisashi Satoh, Satoshi Okabe
    Microbes and environments 35 4 2020年 
    The activity of anaerobic ammonia-oxidizing (anammox) bacteria is considered to depend on cell density; however, this has not yet been confirmed due to the fastidious nature of anammox bacteria (e.g., slow growth, oxygen sensitivity, and rigid aggregate formation). In the present study, the cell density-dependent occurrence of anammox activity (14-15N2 gas production rate) was investigated using planktonic enrichment cultures of Candidatus Brocadia sinica. This activity was detectable when the density of cells was higher than 107‍ ‍cells‍ ‍mL-1 and became stronger with increases in cell density. At the cell densities, the transcription of the BROSI_A1042 and BROSI_A3652 genes, which are potentially involved in the biosynthesis and reception of N-acyl homoserine lactone (AHL), was detectable in Brocadia sinica cells. The presence of AHL molecules in the MBR culture of B. sinica was confirmed by an AHL reporter assay and gas chromatography mass spectrometry analysis. The exogenous addition of the MBR culture extract and AHL molecules (a cocktail of C6, C8, C10, and C12-homoserine lactones) increased the specific 14-15N2 production rate of B. sinica. These results suggest that the specific anammox activity of B. sinica is regulated by AHL-mediated quorum sensing.
  • Kobayashi Kanae, Makabe Akiko, Yano Midori, Oshiki Mamoru, Kindaichi Tomonori, Casciotti Karen L, Okabe Satoshi
    The ISME Journal 13 10 2426 - 2436 2019年10月 [査読有り][通常論文]
  • Kanae Kobayashi, Akiko Makabe, Midori Yano, Mamoru Oshiki, Tomonori Kindaichi, Karen L Casciotti, Satoshi Okabe
    The ISME journal 13 10 2426 - 2436 2019年10月 [査読有り][通常論文]
     
    Natural abundance of stable nitrogen (N) and oxygen (O) isotopes are invaluable biogeochemical tracers for assessing the N transformations in the environment. To fully exploit these tracers, the N and O isotope effects (15ε and 18ε) associated with the respective nitrogen transformation processes must be known. However, the N and O isotope effects of anaerobic ammonium oxidation (anammox), one of the major fixed N sinks and NO3- producers, are not well known. Here, we report the dual N and O isotope effects associated with anammox by three different anammox bacteria including "Ca. Scalindua japonica", a putative marine species, which were measured in continuous enrichment culture experiments. All three anammox species yielded similar N isotope effects of NH4+ oxidation to N2 (15εNH4→N2) ranging from 30.9‰ to 32.7‰ and inverse kinetic isotope effects of NO2- oxidation to NO3- (15εNO2→NO3 = -45.3‰ to -30.1‰). In contrast, 15εNO2→N2 (NO2- reduction to N2) were significantly different among three species, which is probably because individual anammox bacteria species might possess different types of nitrite reductase. We also report the combined O isotope effects for NO2- oxidation (18ENO2→NO3) by anammox bacteria. These obtained dual N and O isotopic effects could provide significant insights into the contribution of anammox bacteria to the fixed N loss and NO2- reoxidation (N recycling) in various natural environments.
  • Zheng Ji, Xiaochang C Wang, Limei Xu, Chongmiao Zhang, Cheng Rong, Andri Taruna Rachmadi, Mohan Amarasiri, Satoshi Okabe, Naoyuki Funamizu, Daisuke Sano
    Pathogens (Basel, Switzerland) 8 4 170 - 170 2019年09月30日 [査読有り][通常論文]
     
    Gastroenteritis viruses in wastewater reclamation systems can pose a major threat to public health. In this study, multiple gastroenteritis viruses were detected from wastewater to estimate the viral contamination sources in a wastewater treatment and reclamation system installed in a suburb of Xi'an city, China. Reverse transcription plus nested or semi-nested PCR, followed by sequencing and phylogenetic analysis, were used for detection and genotyping of noroviruses and rotaviruses. As a result, 91.7% (22/24) of raw sewage samples, 70.8% (17/24) of the wastewater samples treated by anaerobic/anoxic/oxic (A2O) process and 62.5% (15/24) of lake water samples were positive for at least one of target gastroenteritis viruses while all samples collected from membrane bioreactor effluent after free chlorine disinfection were negative. Sequence analyses of the PCR products revealed that epidemiologically minor strains of norovirus GI (GI/14) and GII (GII/13) were frequently detected in the system. Considering virus concentration in the disinfected MBR effluent which is used as the source of lake water is below the detection limit, these results indicate that artificial lake may be contaminated from sources other than the wastewater reclamation system, which may include aerosols, and there is a possible norovirus infection risk by exposure through reclaimed water usage and by onshore winds transporting aerosols containing norovirus.
  • Membrane fouling potentials of an exoelectrogenic fouling-causing bacterium cultured with different external electron acceptors
    So Ishizaki, Rimana Islam Papry, Hiroshi Miyake, Yoko Narita, Satoshi Okabe
    Frontiers in Microbiology 2018年12月 [査読有り][通常論文]
  • Muhammad Ali, Dario Rangel Shaw, Lei Zhang, Mohamed Fauzi Haroon, Yuko Narita, Abdul-Hamid Emwas, Pascal E Saikaly, Satoshi Okabe
    Water research 143 10 - 18 2018年10月15日 [査読有り][通常論文]
     
    Anaerobic ammonium-oxidizing (anammox) bacteria are well known for their aggregation ability. However, very little is known about cell surface physicochemical properties of anammox bacteria and thus their aggregation abilities have not been quantitatively evaluated yet. Here, we investigated the aggregation abilities of three different anammox bacterial species: "Candidatus Brocadia sinica", "Ca. Jettenia caeni" and "Ca. Brocadia sapporoensis". Planktonic free-living enrichment cultures of these three anammox species were harvested from the membrane bioreactors (MBRs). The physicochemical properties (e.g., contact angle, zeta potential, and surface thermodynamics) were analyzed for these anammox bacterial species and used in the extended DLVO theory to understand the force-distance relationship. In addition, their extracellular polymeric substances (EPSs) were characterized by X-ray photoelectron spectroscopy and nuclear magnetic resonance. The results revealed that the "Ca. B. sinica" cells have the most hydrophobic surface and less hydrophilic functional groups in EPS than other anammox strains, suggesting better aggregation capability. Furthermore, aggregate formation and anammox bacterial populations were monitored when planktonic free-living cells were cultured in up-flow column reactors under the same conditions. Rapid development of microbial aggregates was observed with the anammox bacterial population shifts to a dominance of "Ca. B. sinica" in all three reactors. The dominance of "Ca. B. sinica" could be explained by its better aggregation ability and the superior growth kinetic properties (higher growth rate and affinity to nitrite). The superior aggregation ability of "Ca. B. sinica" indicates significant advantages (efficient and rapid start-up of anammox reactors due to better biomass retention as granules and consequently stable performance) in wastewater treatment application.
  • Sunja Cho, Minki Jung, Dongjin Ju, Young-Hee Lee, Kuk Cho, Satoshi Okabe
    Environmental technology 39 19 2503 - 2510 2018年10月 [査読有り][通常論文]
     
    To access the effects of the surface modification and fabric structure of polyethylene (PE) non-woven fabric sheets on retaining the attachment efficiency of anammox biomass, three different non-woven sheets were prepared and inserted in an anammox reactor. The hydrophobic surface modification with 10% KMnO4 and gelatin did not improve the attachment efficiency of the anammox biomass on the surface of the PE non-woven fibers. Densely packed PE-755 having the highest specific surface area to volume ratio (SA/V) (755) retained 221.4 mg biomass per unit sheet, whereas PE-181 having the lowest SA/V (181) retained only 66.4 mg biomass per unit. Accordingly, the volumetric anammox activity of non-woven sheet PE-755 was the highest among the three PE non-woven sheets because of the strong positive relationship between the specific anammox activity and biomass amount (R = 0.835, P < .01). The specific surface area to volume ratio (cm2 cm-3) as well as the bulk density should be considered as important parameters for the selection of non-woven biocarriers for anammox biomass.
  • Andri Taruna Rachmadi, Masaaki Kitajima, Kozo Watanabe, Sakiko Yaegashi, Joeselle Serrana, Arata Nakamura, Toyoko Nakagomi, Osamu Nakagomi, Kazuhiko Katayama, Satoshi Okabe, Daisuke Sano
    Applied and environmental microbiology 84 13 2018年07月01日 [査読有り][通常論文]
     
    Human noroviruses are excreted in feces from infected individuals and included in wastewater. It is critical to remove/inactivate them in wastewater treatment processes, particularly in the disinfection step, before release to aquatic environments. However, the high mutation rates of human noroviruses raise concerns about the emergence of strains that are less susceptible to disinfectants and can survive even after wastewater treatment. This study aimed to demonstrate the strain-dependent susceptibility of norovirus to free chlorine. A population originated from the murine norovirus strain S7-PP3, a surrogate for human noroviruses in environmental testing, was exposed to free chlorine and then propagated in a host cell. This cycle of free chlorine exposure followed by propagation in cells was repeated 10 times, and populations with lower susceptibility to free chlorine were obtained from two independent trials of chlorine exposure cycles. Open reading frame 2 (ORF2) and ORF3 of the murine norovirus genome were analyzed by next-generation sequencing, and a unique nonsynonymous mutation (corresponding to a change from phenylalanine to serine) at nucleotide (nt) 7280 in ORF3, which encodes the minor capsid protein VP2, was found in chlorine-exposed populations from both trials. It was confirmed that all of the clones from the chlorine-treated population had lower susceptibility to free chlorine than those from the control population. These results indicate that exposure to free chlorine and dilution exert different driving forces to form murine norovirus (MNV) quasispecies, and that there is a selective force to form MNV quasispecies under free chlorine exposure.IMPORTANCE This study showed that free chlorine disinfection exerted a selection pressure for murine norovirus (MNV). The strain-dependent viral susceptibility to the disinfectant elucidated in this study highlights the importance of employing less susceptible strains as representative viruses in disinfection tests, because the disinfection rate values obtained from more susceptible strains would be less useful in predicting the virus inactivation efficiency of circulating strains under practical disinfection conditions.
  • Masaaki Kitajima, So Ishizaki, Jeonghwan Jang, Satoshi Ishii, Satoshi Okabe
    Genome announcements 6 23 e00471-18  2018年06月07日 [査読有り][通常論文]
     
    We report here the complete genome sequence of Klebsiella quasipneumoniae strain S05, a bacterium capable of producing membrane fouling-causing soluble substances and capable of respiring on oxygen, nitrate, and an anodic electrode. The genomic information of strain S05 should help predict metabolic pathways associated with these unique biological properties of this bacterium.
  • Akira Hafuka, Hisashi Satoh, Koji Yamada, Masahiro Takahashi, Satoshi Okabe
    Materials (Basel, Switzerland) 11 5 2018年05月16日 [査読有り][通常論文]
     
    We developed an asymmetric fluorescent sensor 1 for Cu2+, based on 4,4-difluoro-4-bora-3a,4a-diaza-s-indacene (BODIPY), by introducing 4-carboxyphenyl and bis(pyridin-2-ylmethyl)amine groups at the 5- and 3-positions, respectively, of the BODIPY core. We then investigated the photophysical and cation-sensing properties of the sensor. BODIPY 1 showed large absorption and fluorescence spectral shifts on binding to Cu2+. The fluorescence peak at 580 nm red-shifted to 620 nm. The binding stoichiometry of BODIPY 1 and Cu2+ was 1:3. The ratio of the fluorescence intensity at 620 nm to that at 580 nm (F620/F580) increased with increasing concentration of Cu2+ (3⁻10 equiv); this enabled ratiometric determination of Cu2+. Although BODIPY 1 showed good selectivity for Cu2+, there was an interfering effect of Fe3+. BODIPY 1 could be used for the naked-eye detection of Cu2+ in a water-containing sample.
  • Rathnayake M L D Rathnayake, Mamoru Oshiki, Satoshi Ishii, Takahiro Segawa, Hisashi Satoh, Satoshi Okabe
    Environmental science & technology 52 10 5744 - 5752 2018年05月15日 [査読有り][通常論文]
     
    Although nitric oxide (NO) emissions from anaerobic ammonium oxidation (anammox)-based processes were reported previously, the NO production pathways are poorly understood. Here, we investigated the NO production pathways in anammox granules in detail by combining 15N-stable isotope tracer experiments with various inhibitors, microsensor measurements, and transcriptome analysis for key genes of NO2- reduction. NO was emitted from the anammox granules, which account for 0.07% of the N2 emission. 15N-stable isotope-tracer experiments indicated that most of the N2 was produced by anammox bacteria, whereas NO was produced from NO2- reduction by anammox and denitrifying bacteria. The NO emission rate was highest at pH 8.0 and accelerated by increasing NH4+ and NO2- concentrations in the culture media. The microsensor analyses showed the in situ NO production rate was highest in the outer layer of the anammox granule where anammox activity was also highest. The detected in situ NO concentrations of up to 2.7 μM were significantly above physiological thresholds known to affect a wide range of microorganisms present in wastewater. Hence, NO likely plays pivotal roles in the microbial interactions in anammox granules, which needs to be further investigated.
  • D. Sano, M. Tazawa, M. Inaba, S. Kadoya, R. Watanabe, T. Miura, M. Kitajima, S. Okabe
    Journal of Applied Microbiology 124 4 1001 - 1007 2018年04月01日 [査読有り][通常論文]
     
    Aims: Cellular responses of an established cell line from human intestinal epithelial cells (INT-407 cells) against poliovirus (PV) infections were investigated in order to find cellular genetic markers for infectious PV detection. Methods and Results: Gene expression profile of INT-407 cells was analysed by DNA microarray technique when cells were infected with poliovirus 1 (PV1) (sabin) at multiplicity of infection of 10−3 and incubated for 12 h. Poliovirus infection significantly altered the gene expressions of two ion channels, KCNJ4 and SCN7A. The expression profile of KCNJ4 gene was further investigated by real-time RT-qPCR, and it was found that KCNJ4 gene was significantly regulated at 24 h postinfection of PV1. Conclusions: KCNJ4 gene, coding a potassium channel protein, is proposed as a cellular genetic marker for infectious PV detection. Significance and Impact of the Study: This is the first study to show the availability of cellular responses to detect infectious PV. The selection of cellular genetic markers for infectious viruses using DNA microarray and RT-qPCR can be applicable for the other enteric viruses.
  • Mohan Amarasiri, Masaaki Kitajima, Akiho Miyamura, Ricardo Santos, Silvia Monteiro, Takayuki Miura, Shinobu Kazama, Satoshi Okabe, Daisuke Sano
    International journal of hygiene and environmental health 221 3 578 - 585 2018年04月 [査読有り][通常論文]
     
    Circulation of human noroviruses in water environments is suspected to be genotype-dependent, but the established primer and probe sets for noroviruses are usually genogroup-specific, which do not allow to compare the genotype-specific properties, such as persistence in water environments and resistance to disinfectants. In this study, quantitative PCR assays were designed for genotype-specific quantification of four epidemiologically important genotypes, GII.3, GII.4, GII.6, and GII.17. Developed assays were tested using norovirus positive stool samples which were previously confirmed to present target genotypes of this study. The results were 100% in accordance with the previous results. Effect of the co-existence of multiple genotypes in a sample on the target genotype quantification was evaluated using composite stool samples and wastewater samples containing multiple genotypes and the presence of non-target genotypes didn't affect the quantification of target genotype. Sensitivity and specificity was 100% for all four assays developed in this study with no cross-reactions between genotypes demonstrating the validity of our assays and their applicability to clinical and environmental samples.
  • Andri Taruna Rachmadi, Masaaki Kitajima, Kozo Watanabe, Satoshi Okabe, Daisuke Sano
    Environmental science & technology 52 5 2434 - 2435 2018年03月06日 [査読有り][通常論文]
  • Electricity generation potential of poultry droppings wastewater in microbial fuel cell using rice husk charcoal electrodes
    Godwin E. Oyiwona, James Chukwuma Ogbonna, Chukwudi Anyanwu, Satoshi Okabe
    Bioresources and Bioprocessing 5 13 1 - 6 2018年03月 [査読有り][通常論文]
  • Insights into the roles of anammox bacteria in post-treatment of anaerobically-treated sewage
    岡部 聡
    Critical Reviews in Environmental Science and Technology 48 6 655 - 684 2018年03月 [査読有り][通常論文]
  • 伊藤 司, 金田一 智則, 押木 守, 岡部 聡
    EICA : journal of EICA : 環境システム計測制御学会誌 22 4 21 - 26 環境システム計測制御学会 2018年 [査読有り][通常論文]
  • Qian Zhang, Gabriel A Al-Ghalith, Mayumi Kobayashi, Takahiro Segawa, Mitsuto Maeda, Satoshi Okabe, Dan Knights, Satoshi Ishii
    Frontiers in microbiology 9 2201 - 2201 2018年 [査読有り][通常論文]
     
    Current approach to identify sources of human pathogens is largely dependent on the cultivation and isolation of target bacteria. For rapid pathogen source identification, culture-independent strain typing method is necessary. In this study, we designed new primer set that broadly covers flaA short variable region (SVR) of various Campylobacter species, and applied the flaA SVR sequencing method to examine the diversity of Campylobacter spp. in geese fecal samples (n = 16) with and without bacteria cultivation. Twenty-three Campylobacter strains isolated from the 16 geese fecal samples were grouped similarly by conventional flaA restriction fragment length polymorphism (RFLP) method and by the flaA SVR sequencing method, but higher discriminant power was observed in the flaA SVR sequencing approach. For culture-independent flaA SVR sequencing analysis, we developed and optimized the sequence data analysis pipeline to identify as many genotypes as possible, while minimizing the detection of genotypes generated by sequencing errors. By using this pipeline, 51,629 high-quality flaA sequence reads were clustered into 16 operational taxonomic units (=genotypes) by using 98% sequence similarity and >50 sequence duplicates. Almost all flaA genotypes obtained by culture-dependent method were also identified by culture-independent flaA SVR MiSeq sequencing method. In addition, more flaA genotypes were identified probably due to high throughput nature of the MiSeq sequencing. These results suggest that the flaA SVR sequencing could be used to analyze the diversity of Campylobacter spp. without bacteria isolation. This method is promising to rapidly identify potential sources of Campylobacter pathogens.
  • Lei Zhang, Satoshi Okabe
    Water research 127 204 - 210 2017年12月15日 [査読有り][通常論文]
     
    Despite that anaerobic ammonium oxidizing (anammox) bacteria are key players in the global nitrogen cycle, no pure cultures are still available. Planktonic cell culture with high purity is, therefore, essential for physiological and biochemical studies of anammox bacteria. However, development of such planktonic cell cultures requires an enormous amount of time and effort. Here we developed a novel rapid method for cultivating free-living planktonic anammox cells. First, anammox granules were physically dispersed, immobilized in 6% polyvinyl alcohol-4% sodium alginate (PVA-SA) gel beads, and then pre-cultured in an up-flow column reactor. Anammox bacteria grew rapidly as loosely flocculated micro-clusters in the gel beads. After 18 days of pre-cultivation, mature gel beads were harvested, physically dispersed by vortex and inoculated into a membrane bioreactor (MBR). The MBR was then continuously operated at a low nitrogen loading rate (<0.9 kg-TN m-3 d-1). After 17 days of operation, active free-living planktonic anammox cells with purity >95% were successfully developed in the MBR. Total culture time (gel beads and MBR) to accomplish free-living planktonic anammox cells was only 35 days, which was significantly shorter than the previous reports. This new cultivation technique could greatly facilitate various microbial, physiological and biochemical studies of anammox bacteria.
  • Toshikazu Fukushima, Hiroe Hara-Yamamura, Koji Nakashima, Lea Chua Tan, Satoshi Okabe
    Chemosphere 188 312 - 319 2017年12月 [査読有り][通常論文]
     
    Wastewater effluents contain a significant number of toxic contaminants, which, even at low concentrations, display a wide variety of toxic actions. In this study, we developed a multiple-endpoints gene alteration-based (MEGA) assay, a real-time PCR-based transcriptomic analysis, to assess the water quality of wastewater effluents for human health risk assessment and management. Twenty-one genes from the human hepatoblastoma cell line (HepG2), covering the basic health-relevant stress responses such as response to xenobiotics, genotoxicity, and cytotoxicity, were selected and incorporated into the MEGA assay. The genes related to the p53-mediated DNA damage response and cytochrome P450 were selected as markers for genotoxicity and response to xenobiotics, respectively. Additionally, the genes that were dose-dependently regulated by exposure to the wastewater effluents were chosen as markers for cytotoxicity. The alterations in the expression of an individual gene, induced by exposure to the wastewater effluents, were evaluated by real-time PCR and the results were validated by genotoxicity (e.g., comet assay) and cell-based cytotoxicity tests. In summary, the MEGA assay is a real-time PCR-based assay that targets cellular responses to contaminants present in wastewater effluents at the transcriptional level; it is rapid, cost-effective, and high-throughput and can thus complement any chemical analysis for water quality assessment and management.
  • Lei Zhang, Yuko Narita, Lin Gao, Muhammad Ali, Mamoru Oshiki, Satoshi Ishii, Satoshi Okabe
    Water research 125 249 - 258 2017年11月15日 [査読有り][通常論文]
     
    Phylogenetically diverse anammox bacteria have been detected in most of anoxic natural and engineered ecosystems and thus regarded as key players in the global nitrogen cycle. However, ecological niche differentiation of anammox bacteria remains unresolved despite its ecological and practical importance. In this study, the microbial competitions for a common substrate (nitrite) among three anammox species (i.e. "Candidatus Brocadia sinica", "Candidatus Jettenia caeni" and "Candidatus Kuenenia stuttgartiensis") were systematically investigated in nitrite-limited gel-immobilized column reactors (GICR) and membrane bioreactors (MBRs) under different nitrogen loading rates (NLRs). 16 S rRNA gene-based population dynamics revealed that "Ca. J. caeni" could proliferate only at low NLRs, whereas "Ca. B. sinica" outcompeted other two species at higher NLRs in both types of reactors. Furthermore, FISH analysis revealed that "Ca. J. caeni" was mainly present as spherical microclusters at the inner part (low NO2- environment), whereas "Ca. B. sinica" was present throughout the gel beads and granules. This spatial distribution supports the outcomes of the competition experiments. However, the successful competition of "Ca. J. caeni" at low NLR could not be explained with the Monod model probably due to inaccuracy of kinetic parameters such as half saturation constant (Ks) for nitrite and a difference in the maintenance rate (m). In addition, the growth of "Ca. K. stuttgartiensis" could not be observed in any experimental conditions, suggesting possible unknown factor(s) is missing. Taken together, NLR was one of factors determining ecological niche differentiation of "Ca. B. sinica" and "Ca. J. caeni".
  • Toshihiro Ito, Masaaki Kitajima, Tsuyoshi Kato, Satoshi Ishii, Takahiro Segawa, Satoshi Okabe, Daisuke Sano
    Water research 125 438 - 448 2017年11月15日 [査読有り][通常論文]
     
    Multiple-barriers are widely employed for managing microbial risks in water reuse, in which different types of wastewater treatment units (biological treatment, disinfection, etc.) and health protection measures (use of personal protective gear, vegetable washing, etc.) are combined to achieve a performance target value of log10 reduction (LR) of viruses. The LR virus target value needs to be calculated based on the data obtained from monitoring the viruses of concern and the water reuse scheme in the context of the countries/regions where water reuse is implemented. In this study, we calculated the virus LR target values under two exposure scenarios for reclaimed wastewater irrigation in Japan, using the concentrations of indigenous viruses in untreated wastewater and a defined tolerable annual disease burden (10-4 or 10-6 disability-adjusted life years per person per year (DALYpppy)). Three genogroups of norovirus (norovirus genogroup I (NoV GI), geogroup II (NoV GII), and genogroup IV (NoV GIV)) in untreated wastewater were quantified as model viruses using reverse transcription-microfluidic quantitative PCR, and only NoV GII was present in quantifiable concentration. The probabilistic distribution of NoV GII concentration in untreated wastewater was then estimated from its concentration dataset, and used to calculate the LR target values of NoV GII for wastewater treatment. When an accidental ingestion of reclaimed wastewater by Japanese farmers was assumed, the NoV GII LR target values corresponding to the tolerable annual disease burden of 10-6 DALYpppy were 3.2, 4.4, and 5.7 at 95, 99, and 99.9%tile, respectively. These percentile values, defined as "reliability," represent the cumulative probability of NoV GII concentration distribution in untreated wastewater below the corresponding tolerable annual disease burden after wastewater reclamation. An approximate 1-log10 difference of LR target values was observed between 10-4 and 10-6 DALYpppy. The LR target values were influenced mostly by the change in the logarithmic standard deviation (SD) values of NoV GII concentration in untreated wastewater and the reliability values, which highlights the importance of accurately determining the probabilistic distribution of reference virus concentrations in source water for water reuse.
  • Hisashi Satoh, Wasala M K R T W Bandara, Manabu Sasakawa, Yoshihito Nakahara, Masahiro Takahashi, Satoshi Okabe
    Bioresource technology 244 Pt 1 768 - 775 2017年11月 [査読有り][通常論文]
     
    A hollow fiber degassing membrane (DM) was applied to enhance organic matter degradation and methane gas production of anaerobic granular sludge process by reducing the dissolved hydrogen gas (D-H2) concentration in the liquid phase. DM was installed in the bench-scale anaerobic granular sludge reactors and D-H2 was removed through DM using a vacuum pump. Degasification improved the organic matter degradation efficiency to 79% while the efficiency was 62% without degasification at 12,000mgL-1 of the influent T-COD concentration. Measurement of D-H2 concentrations in the liquid phase confirmed that D-H2 was removed by degasification. Furthermore, the effect of acetate concentrations on the organic matter degradation efficiency was investigated. At acetate concentrations above 3gL-1, organic matter degradation deteriorated. Degasification enhanced the propionate and acetate degradation. These results suggest that degasification reduced D-H2 concentration and volatile fatty acids concentrations, prevented pH drop, and subsequent enhanced organic matter degradation.
  • Akira Hafuka, Akiyoshi Takitani, Hiroko Suzuki, Takuya Iwabuchi, Masahiro Takahashi, Satoshi Okabe, Hisashi Satoh
    Sensors (Basel, Switzerland) 17 10 2017年10月09日 [査読有り][通常論文]
     
    Simple analytical methods are needed for determining the cadmium (Cd) content of brown rice samples. In the present study, we developed a new analytical procedure consisting of the digestion of rice using HCl, Cd purification using anion exchange resin, and then determining the Cd content using fluorescence spectroscopy. Digestion with 0.1 M HCl for 10 min at room temperature was sufficient to extract Cd from the ground rice samples. The Cd in the extract was successfully purified in preference to other metals using Dowex 1X8 chloride form resin. Low concentrations of Cd in the eluate could be determined using fluorescence spectroscopy with a fluoroionophore. Overall, the actual limit of quantification value for the Cd content in rice was about 0.1 mg-Cd/kg-rice, which was sufficiently low compared with the regulatory value (0.4 mg-Cd/kg-rice) given by the Codex Alimentarius Commission. We analyzed authentic brown rice samples using our new analytical procedure and the results agreed well with those determined using inductively coupled plasma optical emission spectrometry (ICP-OES). Since the fluoroionophore recognized Zn2+ and Hg2+ as well as Cd2+, a sample containing high concentration of Zn2+ or Hg2+ might cause a false positive result.
  • Mamoru Oshiki, Keisuke Mizuto, Zen-Ichiro Kimura, Tomonori Kindaichi, Hisashi Satoh, Satoshi Okabe
    Environmental microbiology reports 9 5 550 - 561 2017年10月 [査読有り][通常論文]
     
    Anaerobic ammonium-oxidizing (anammox) bacteria affiliated with the genus 'Candidatus Scalindua' are responsible for significant nitrogen loss in oceans, and thus their ecophysiology is of great interest. Here, we enriched a marine anammox bacterium, 'Ca. S. japonica' from a Hiroshima bay sediment in Japan, and comparative genomic and proteomic analyses of 'Ca. S. japonica' were conducted. Sequence of the 4.81-Mb genome containing 4019 coding regions of genes (CDSs) composed of 47 contigs was determined. In the proteome, 1762 out of 4019 CDSs in the 'Ca. S. japonica' genome were detected. Based on the genomic and proteomic data, the core anammox process and carbon fixation of 'Ca. S. japonica' were further investigated. Additionally, the present study provides the first detailed insights into the genetic background responsible for iron acquisition and menaquinone biosynthesis in anammox bacterial cells. Comparative analysis of the 'Ca. Scalindua' genomes revealed that the 1502 genes found in the 'Ca. S. japonica' genome were not present in the 'Ca. S. profunda' and 'Ca. S. rubra' genomes, showing a high genomic diversity. This result may reflect a high phylogenetic diversity of the genus 'Ca. Scalindua'.
  • Yuko Narita, Lei Zhang, Zen-Ichiro Kimura, Muhammad Ali, Takao Fujii, Satoshi Okabe
    Systematic and applied microbiology 40 7 448 - 457 2017年10月 [査読有り][通常論文]
     
    We successfully enriched a novel anaerobic ammonium-oxidizing (anammox) bacterium affiliated with the genus 'Candidatus Brocadia' with high purity (>90%) in a membrane bioreactor (MBR). The enriched bacterium was distantly related to the hitherto characterized 'Ca. Brocadia fulgida' and 'Ca. Brocadia sinica' with 96% and 93% of 16S ribosomal RNA gene sequence identity, respectively. The bacterium exhibited the common structural features of anammox bacteria and produced hydrazine in the presence of hydroxylamine under anoxic conditions. The temperature range of anammox activity was 20-45°C with a maximum activity at 37°C. The maximum specific growth rate (μmax) was 0.0082h-1 at 37°C, corresponding to a doubling time of 3.5 days. The half-saturation constant (KS) for nitrite was 5±2.5μM. The anammox activity was inhibited by nitrite (IC50=11.6mM) but not by formate and acetate. The major respiratory quinone was identified to be menaquinone-7 (MK-7). The enriched anammox bacterium shared nearly half of genes with 'Ca. Brocadia sinica' and 'Ca. Brocadia fulgida'. The enriched bacterium showed all known physiological characteristics of anammox bacteria and can be distinguished from the close relatives by its 16S rRNA gene sequence. Therefore, we proposed the name 'Ca. Brocadia sapporoensis' sp. nov.
  • Mohan Amarasiri, Masaaki Kitajima, Thanh H Nguyen, Satoshi Okabe, Daisuke Sano
    Water research 121 258 - 269 2017年09月15日 [査読有り][通常論文]
     
    The multiple-barrier concept is widely employed in international and domestic guidelines for wastewater reclamation and reuse for microbiological risk management, in which a wastewater reclamation system is designed to achieve guideline values of the performance target of microbe reduction. Enteric viruses are one of the pathogens for which the target reduction values are stipulated in guidelines, but frequent monitoring to validate human virus removal efficacy is challenging in a daily operation due to the cumbersome procedures for virus quantification in wastewater. Bacteriophages have been the first choice surrogate for this task, because of the well-characterized nature of strains and the presence of established protocols for quantification. Here, we performed a meta-analysis to calculate the average log10 reduction values (LRVs) of somatic coliphages, F-specific phages, MS2 coliphage and T4 phage by membrane bioreactor, activated sludge, constructed wetlands, pond systems, microfiltration and ultrafiltration. The calculated LRVs of bacteriophages were then compared with reported human enteric virus LRVs. MS2 coliphage LRVs in MBR processes were shown to be lower than those of norovirus GII and enterovirus, suggesting it as a possible validation and operational monitoring tool. The other bacteriophages provided higher LRVs compared to human viruses. The data sets on LRVs of human viruses and bacteriophages are scarce except for MBR and conventional activated sludge processes, which highlights the necessity of investigating LRVs of human viruses and bacteriophages in multiple treatment unit processes.
  • So Ishizaki, Ryoichi Sugiyama, Satoshi Okabe
    Scientific reports 7 1 8482 - 8482 2017年08月16日 [査読有り][通常論文]
     
    Membrane fouling still remains a major obstacle for wider applications of membrane bioreactor (MBR), which is mainly caused by soluble microbial products (SMP). Identification of key bacteria responsible for SMP production is essential for mitigation of membrane fouling. Here, we investigated the effect of microbial interaction on membrane fouling. We measured the membrane fouling potentials of 13 bacterial strains isolated from a pilot-scale MBR treating domestic wastewater when they were cultivated as single-culture and co-culture. We found that fouling-causing bacteria (FCB) displayed much higher fouling potential when co-cultured even with non-FCB and mixed population (activated sludge). In particular, the fouling potential of strain S26, one of FCB, increased 26.8 times when cultivated with strain S22 (fouling-enhancing bacteria, FEB). The secretion of N-octanoyl-L-homoserine lactone (C8-HSL) was increased by co-cultivating S22 and S26 as compared with cultivating as single culture, which stimulated the production of fouling-causing SMP by S26 and consequently resulted in severe membrane fouling. This result suggests that AHL-mediated quorum-sensing (QS) regulatory system was involved in secretion of fouling-causing SMP.
  • Lei Zhang, Yuko Narita, Lin Gao, Muhammad Ali, Mamoru Oshiki, Satoshi Okabe
    Water research 116 296 - 303 2017年06月01日 [査読有り][通常論文]
     
    Anammox bacteria have long been considered to be slow-growing bacteria. However, it has recently been reported that they could grow much faster than previously thought when they were cultivated in a membrane bioreactor (MBR) with a step-wise decrease in the solid retention time (SRT). Here, we reevaluated the maximum specific growth rates (μmax) of three phylogenetically distant anammox bacterial species (i.e. "Ca. Brocadia sinica", "Ca. Jettenia caeni" and "Ca. Scalindua sp.") by directly measuring 16S rRNA gene copy numbers using newly developed quantitative polymerase chain reaction (qPCR) assays. When free-living planktonic "Ca. B. sinica" and "Ca. J. caeni" cells were immobilized in polyvinyl alcohol (PVA) and sodium alginate (SA) gel beads and cultivated in an up-flow column reactor with high substrate loading rates at 37 °C, the μmax were determined to be 0.33 ± 0.02 d-1 and 0.18 d-1 (corresponding doubling time of 2.1 day and 3.9 day) from the exponential increases in 16S rRNA genes copy numbers, respectively. These values were faster than the fastest growth rates reported for these species so far. The cultivation of anammox bacteria in gel beads was achieved less than one month without special cultivation method and selection pressure, and the exponential increase in 16S rRNA gene numbers was directly measured by qPCR with high reproducibility; therefore, the resulting μmax values were considered accurate. Taken together, the fast growth is, therefore, considered to be an intrinsic kinetic property of anammox bacteria.
  • Fumika Nishino, Melbert Jeem, Lihua Zhang, Kazumasa Okamoto, Satoshi Okabe, Seiichi Watanabe
    Scientific reports 7 1 1063 - 1063 2017年04月21日 [査読有り][通常論文]
     
    We report the fabrication of flower-like CuO nanostructured surfaces via submerged photo-synthesis of crystallites (SPSC), which requires only UV illumination in neutral water. In this paper, we discuss the reaction mechanism of the photochemical formation of the SPSC-fabricated CuO nanostructures in detail based on surface microstructural analyses and a radiation-chemical consideration with additional gamma-ray irradiation. Since the SPSC method for surface nanostructural fabrication can work at low temperatures at atmospheric pressure without using harmful substances, it is a potential fabrication method for green nanotechnology applications. In this vein, the antibacterial activity of the nano-flowered CuO surfaces was tested against Gram-positive (Staphylococcus aureus) bacteria and Gram-negative (Escherichia coli K12) bacteria, and the results demonstrate that the nano-flowered CuO nanostructures act as an effective antimicrobial agent.
  • Yosuke Tashiro, Hiroaki Eida, Satoshi Ishii, Hiroyuki Futamata, Satoshi Okabe
    Microbes and environments 32 1 40 - 46 2017年03月31日 [査読有り][通常論文]
     
    A conjugative F plasmid induces mature biofilm formation by Escherichia coli by promoting F-pili-mediated cell-cell interactions and increasing the expression of biofilm-related genes. We herein demonstrated another function for the F plasmid in E. coli biofilms; it contributes to the emergence of genetic and phenotypic variations by spontaneous mutations. Small colony variants (SCVs) were more frequently generated in a continuous flow-cell biofilm than in the planktonic state of E. coli harboring the F plasmid. E. coli SCVs represented typical phenotypic changes such as slower growth, less biofilm formation, and greater resistance to aminoglycoside antibiotics than the parent strain. Genomic and complementation analyses indicated that the small colony phenotype was caused by the insertion of Tn1000, which was originally localized in the F plasmid, into the hemB gene. Furthermore, the Tn1000 insertion was removed from hemB in the revertant, which showed a normal colony phenotype. This study revealed that the F plasmid has the potential to increase genetic variations not only by horizontal gene transfer via F pili, but also by site-specific recombination within a single cell.
  • Godwin E Oyiwona, James Ogbonna, Chukwudi Uzoma Anyanwu, So Ishizaki, Zen-Ichiro Kimura, Satoshi Okabe
    Biotechnology letters 39 2 253 - 259 2017年02月 [査読有り][通常論文]
     
    OBJECTIVE: To investigate a syntrophic interaction between Geobacter sulfurreducens and hydrogenotrophic methanogens in sludge-inoculated microbial fuel cell (MFC) systems running on glucose with an improved electron recovery at the anode. RESULTS: The presence of archaea in MFC reduces Coulombic efficiency (CE) due to their electron scavenging capability but, here, we demonstrate that a syntrophic interaction can occur between G. sulfurreducens and hydrogenotrophic methanogens via interspecies H2 transfer with improvement in CE and power density. The addition of the methanogenesis inhibitor, 2-bromoethanesulfonate (BES), resulted in the reduction in power density from 5.29 to 2 W/m3, and then gradually increased to the peak value of 5.5 W/m3 when BES addition was stopped. CONCLUSION: Reduction of H2 partial pressure by archaea is an efficient approach in improving power output in a glucose-fed MFC system using Geobacter sp. as an inoculum.
  • Hisashi Satoh, Yuji Miyazaki, Shou Taniuchi, Mamoru Oshiki, Rathnayake M L D Rathnayake, Masahiro Takahashi, Satoshi Okabe
    Analytical sciences : the international journal of the Japan Society for Analytical Chemistry 33 7 825 - 830 2017年 [査読有り][通常論文]
     
    An ionophore-doped sensing membrane phosphate (PO4) microsensor based on bis(dibromophenylstannyl)methane (Bis microsensor) is described. The Bis microsensor showed a Nernstian response. The response of the Bis microsensor was log-linear down to a monohydrogen phosphate ion (HPO42-) concentration of 0.5 μM (corresponding to 1.0 μM of orthophosphate at pH 7.2), whereas the detection limit of PO4-microsensors based on trialkyl/aryltin chloride was 50 μM of HPO42-. The Bis microsensor showed excellent selectivity for HPO42- against nitrite, nitrate, chloride, bicarbonate and sulfate, as compared with PO4 microsensors based on trialkyl/aryltin chloride. Dissolved oxygen, which is known to interfere with the response of a previously developed cobalt-based potentiometric solid-state PO4 microsensor, had no effect on the response of the ionophore-doped sensing membrane-type microsensors described herein. Only OH- (i.e., pH) interfered with the ionophore-doped sensing membrane-type microsensors.
  • Muhammad Ali, Mohamed Fauzi Haroon, Yuko Narita, Lei Zhang, Dario Rangel Shaw, Satoshi Okabe, Pascal E Saikaly
    Genome announcements 4 6 e01377-16  2016年12月08日 [査読有り][通常論文]
     
    The anaerobic ammonium-oxidizing (anammox) bacterium "Candidatus Brocadia sp. 40" demonstrated the fastest growth rate compared to others in this taxon. Here, we report the 2.93-Mb draft genome sequence of this bacterium, which has 2,565 gene-coding regions, 41 tRNAs, and a single rrn operon.
  • Toshihiro Ito, Tsuyoshi Kato, Makoto Hasegawa, Hiroyuki Katayama, Satoshi Ishii, Satoshi Okabe, Daisuke Sano
    Journal of water and health 14 6 879 - 889 2016年12月 [査読有り][通常論文]
     
    The virus reduction efficiency of each unit process is commonly determined based on the ratio of virus concentration in influent to that in effluent of a unit, but the virus concentration in wastewater has often fallen below the analytical quantification limit, which does not allow us to calculate the concentration ratio at each sampling event. In this study, left-censored datasets of norovirus (genogroup I and II), and adenovirus were used to calculate the virus reduction efficiency in unit processes of secondary biological treatment and chlorine disinfection. Virus concentration in influent, effluent from the secondary treatment, and chlorine-disinfected effluent of four municipal wastewater treatment plants were analyzed by a quantitative polymerase chain reaction (PCR) approach, and the probabilistic distributions of log reduction (LR) were estimated by a Bayesian estimation algorithm. The mean values of LR in the secondary treatment units ranged from 0.9 and 2.2, whereas those in the free chlorine disinfection units were from -0.1 and 0.5. The LR value in the secondary treatment was virus type and unit process dependent, which raised the importance for accumulating the data of virus LR values applicable to the multiple-barrier system, which is a global concept of microbial risk management in wastewater reclamation and reuse.
  • Muhammad Ali, Rathnayake M L D Rathnayake, Lei Zhang, Satoshi Ishii, Tomonori Kindaichi, Hisashi Satoh, Sakae Toyoda, Naohiro Yoshida, Satoshi Okabe
    Water research 102 147 - 157 2016年10月01日 [査読有り][通常論文]
     
    Nitrous oxide (N2O) production pathway in a signal-stage nitritation-anammox sequencing batch reactor (SBR) was investigated based on a multilateral approach including real-time N2O monitoring, N2O isotopic composition analysis, and in-situ analyses of spatial distribution of N2O production rate and microbial populations in granular biomass. N2O emission rate was high in the initial phase of the operation cycle and gradually decreased with decreasing NH4(+) concentration. The average emission of N2O was 0.98 ± 0.42% and 1.35 ± 0.72% of the incoming nitrogen load and removed nitrogen, respectively. The N2O isotopic composition analysis revealed that N2O was produced via NH2OH oxidation and NO2(-) reduction pathways equally, although there is an unknown influence from N2O reduction and/or anammox N2O production. However, the N2O isotopomer analysis could not discriminate the relative contribution of nitrifier denitrification and heterotrophic denitrification in the NO2(-) reduction pathway. Various in-situ techniques (e.g. microsensor measurements and FISH (fluorescent in-situ hybridization) analysis) were therefore applied to further identify N2O producers. Microsensor measurements revealed that approximately 70% of N2O was produced in the oxic surface zone, where nitrifiers were predominantly localized. Thus, NH2OH oxidation and NO2 reduction by nitrifiers (nitrifier-denitrification) could be responsible for the N2O production in the oxic zone. The rest of N2O (ca. 30%) was produced in the anammox bacteria-dominated anoxic zone, probably suggesting that NO2(-) reduction by coexisting putative heterotrophic denitrifiers and some other unknown pathway(s) including the possibility of anammox process account for the anaerobic N2O production. Further study is required to identify the anaerobic N2O production pathways. Our multilateral approach can be useful to quantitatively examine the relative contributions of N2O production pathways. Good understanding of the key N2O production pathways is essential to establish a strategy to mitigate N2O emission from biological nitrogen removal processes.
  • Nowaki Hijikata, Rui Tezuka, Shinobu Kazama, Masahiro Otaki, Ken Ushijima, Ryusei Ito, Satoshi Okabe, Daisuke Sano, Naoyuki Funamizu
    Journal of environmental management 181 721 - 727 2016年10月01日 [査読有り][通常論文]
     
    In the present study, the bactericidal and virucidal mechanisms in the alkaline disinfection of compost with calcium lime and ash were investigated. Two indicator microorganisms, Escherichia coli and MS2 coliphage, were used as surrogates for enteric pathogens. The alkaline-treated compost with calcium oxide (CaO) or ash resulted primarily in damage to the outer membrane and enzyme activities of E. coli. The alkaline treatment of compost also led to the infectivity loss of the coliphage because of the partial capsid damage and RNA exteriorization due to a raised pH, which is proportional to the amount of alkaline agents added. These results indicate that the alkaline treatment of compost using calcium oxide and ash is effective and can contribute to the safe usage of compost from a mixing type dry toilet.
  • So Ishizaki, Kotaro Terada, Hiroshi Miyake, Satoshi Okabe
    Environmental science & technology 50 17 9515 - 23 2016年09月06日 [査読有り][通常論文]
     
    Microbial fuel cells (MFCs) have recently been integrated with membrane bioreactors (MBRs) for wastewater treatment and energy recovery. However, the impact of integration of the two reactors on membrane fouling of MBR has not been reported yet. In this study, MFCs equipped with different external resistances (1-10 000 ohm) were operated, and membrane-fouling potentials of the MFC anode effluents were directly measured to study the impact of anodic respiration by exoelectrogens on membrane fouling. It was found that although the COD removal efficiency was comparable, the fouling potential was significantly reduced due to less production of biopolymer (a major foulant) in MFCs equipped with lower external resistance (i.e., with higher current generation) as compared with aerobic respiration. Furthermore, it was confirmed that Geobacter sulfurreducens strain PCA, a dominant exoelectrogen in anode biofilms of MFCs in this study, produced less biopolymer under anodic respiration condition than fumarate (anaerobic) respiration condition, resulting in lower membrane-fouling potential. Taken together, anodic respiration can mitigate membrane fouling of MBR due to lower biopolymer production, suggesting that development of an electrode-assisted MBR (e-MBR) without aeration is feasible.
  • So Ishizaki, Toshikazu Fukushima, Satoshi Ishii, Satoshi Okabe
    Water research 100 448 - 457 2016年09月01日 [査読有り][通常論文]
     
    Membrane fouling remains a major challenge for wider application of membrane bioreactors (MBRs) to wastewater treatment. Membrane fouling is mainly caused by microorganisms and their excreted microbial products. For development of more effective control strategies, it is important to identify and characterize the microorganisms that are responsible for membrane fouling. In this study, 41 bacterial strains were isolated from fouled microfiltration membranes in a pilot-scale MBR treating real municipal wastewater, and their membrane fouling potentials were directly measured using bench-scale cross-flow membrane filtration systems (CFMFSs) and related to their cellular properties. It was found that the fouling potential was highly strain dependent, suggesting that bacterial identification at the strain level is essential to identify key fouling-causing bacteria (FCB). The FCB showed some common cellular properties. The most prominent feature of FCB was that they formed convex colonies having swollen podgy shape and smooth lustrous surfaces with high water, hydrophilic organic matter and carbohydrate content. However, general and rigid biofilm formation potential as determined by microtiter plates and cell surface properties (i.e., hydrophobicity and surface charge) did not correlate with the fouling potential in this study. These results suggest that the fouling potential should be directly evaluated under filtration conditions, and the colony water content could be a useful indicator to identify the FCB.
  • Mamoru Oshiki, Hisashi Satoh, Satoshi Okabe
    Environmental microbiology 18 9 2784 - 96 2016年09月 [査読有り][通常論文]
     
    Anaerobic ammonium oxidation (anammox) is a microbial process in which NH4 (+) is oxidized to N2 gas with NO2 (-) as an electron acceptor. The anammox process is mediated by bacterial members affiliated with the phylum Planctomycetes, which are ubiquitously detected from anoxic natural and man-made ecosystems and a key player in the global nitrogen cycle. In the past two decades, phylogenetically different anammox bacteria have been recognized in natural and synthetic ecosystems (i.e. 'Candidatus Kuenenia', 'Candidatus Brocadia', 'Candidatus Jettenia', 'Candidatus Anammoxoglobus' and 'Candidatus Scalindua' genera), and the geographic distributions of these anammox bacteria indicate that they have genus-specific or species-specific habitats. Recently, we revealed the physiological characteristics of 'Ca. Jettenia' in addition to 'Ca. Kuenenia', 'Ca. Brocadia' and 'Ca. Scalindua', and, as a result, it is possible to compare the physiological characteristics of the anammox bacteria and discuss their niche partitioning. Therefore, we summarize the current knowledge of anammox bacterial ecology and physiology in this review to assess the potential ecological niche partitioning of anammox bacteria in natural and synthetic ecosystems.
  • Mamoru Oshiki, Muhammad Ali, Kaori Shinyako-Hata, Hisashi Satoh, Satoshi Okabe
    Environmental microbiology 18 9 3133 - 43 2016年09月 [査読有り][通常論文]
     
    Although metabolic pathways and associated enzymes of anaerobic ammonium oxidation (anammox) of 'Ca. Kuenenia stuttgartiensis' have been studied, those of other anammox bacteria are still poorly understood. NO2- reduction to NO is considered to be the first step in the anammox metabolism of 'Ca. K. stuttgartiensis', however, 'Ca. Brocadia' lacks the genes that encode canonical NO-forming nitrite reductases (NirS or NirK) in its genome, which is different from 'Ca. K. stuttgartiensis'. Here, we studied the anammox metabolism of 'Ca. Brocadia sinica'. (15) N-tracer experiments demonstrated that 'Ca. B. sinica' cells could reduce NO2- to NH2 OH, instead of NO, with as yet unidentified nitrite reductase(s). Furthermore, N2 H4 synthesis, downstream reaction of NO2- reduction, was investigated using a purified 'Ca. B. sinica' hydrazine synthase (Hzs) and intact cells. Both the 'Ca. B. sinica' Hzs and cells utilized NH2 OH and NH4+, but not NO and NH4+, for N2 H4 synthesis and further oxidized N2 H4 to N2 gas. Taken together, the metabolic pathway of 'Ca. B. sinica' is NH2 OH-dependent and different from the one of 'Ca. K. stuttgartiensis', indicating metabolic diversity of anammox bacteria.
  • Satoshi Ishii, Kazuki Joikai, Shigeto Otsuka, Keishi Senoo, Satoshi Okabe
    MICROBES AND ENVIRONMENTS 31 3 293 - 298 2016年09月 [査読有り][通常論文]
     
    Pseudogulbenkiania is a relatively recently characterized genus within the order Neisseriales, class Betaproteobacteria. This genus contains several strains that are capable of anaerobic, nitrate-dependent Fe(II) oxidation (NDFO), a geochemically important reaction for nitrogen and iron cycles. In the present study, we examined denitrification functional gene diversities within this genus, and clarified whether other Pseudogulbenkiania sp. strains perform denitrification and NDFO. Seventy strains were analyzed, including two type strains, a well-characterized NDFO strain, and 67 denitrifying strains isolated from various rice paddy fields and rice-soybean rotation fields in Japan. We also attempted to identify the genes responsible for NDFO by mutagenesis. Our comprehensive analysis showed that all Pseudogulbenkiania strains tested performed denitrification and NDFO; however, we were unable to obtain NDFO-deficient denitrifying mutants in our mutagenesis experiment. This result suggests that Fe(II) oxidation in these strains is not enzymatic, but is caused by reactive N-species that are formed during nitrate reduction. Based on the results of the comparative genome analysis among Pseudogulbenkiania sp. strains, we identified low sequence similarity within the nos gene as well as different gene arrangements within the nos gene cluster, suggesting that nos genes were horizontally transferred. Since Pseudogulbenkiania sp. strains have been isolated from various locations around the world, their denitrification and NDFO abilities may contribute significantly to nitrogen and iron biogeochemical cycles.
  • Satoshi Ishii, Mohan Amarasiri, Satoshi Hashiba, Peiyi Yang, Satoshi Okabe, Daisuke Sano
    Genome announcements 4 4 e00893 - 16 2016年08月25日 [査読有り][通常論文]
     
    Enterobacter sp. strain SENG-6, isolated from healthy human feces, produces histo-blood group antigen (HBGA)-like substances that can bind with human noroviruses. Based on the genome sequence analysis, strain SENG-6 belongs to the species Enterobacter cloacae The genome sequence of this strain should help identify genes associated with the production of HBGA-like substances.
  • Shuji Ozawa, Satoshi Okabe, Satoshi Ishii
    Foodborne pathogens and disease 13 8 456 - 61 2016年08月 [査読有り][通常論文]
     
    Contamination of food and water with pathogenic bacteria is of concern. Although culture-independent detection and quantification of pathogens is useful, isolation of pathogenic bacteria is still important when identifying the sources of pathogens. Here, we report the use of flow cytometry (FCM) and fluorescence-activated cell sorting (FACS) to specifically detect and isolate individual Escherichia coli O157:H7 cells from water samples. When present at >10 cells/mL water, target pathogen was specifically detected and isolated. The FACS-sorted E. coli O157:H7 population reflected the original population diversity, in contrast to the populations obtained by immunomagnetic separation. Relative abundance of multiple pathogenic strains is important when performing source-tracking studies; therefore, single-cell isolation with FCM-FACS can be a useful tool to obtain pathogenic bacteria for source tracking purpose.
  • 伊藤寿宏, 押木守, 小林直央, 加藤毅, 瀬川高弘, 幡本将史, 山口隆司, 原田秀樹, 北島正章, 岡部聡, 佐野大輔
    土木学会論文集 72 7 305 - 313 2016年07月 [査読有り][通常論文]
  • Mohan Amarasiri, Satoshi Hashiba, Takayuki Miura, Toyoko Nakagomi, Osamu Nakagomi, Satoshi Ishii, Satoshi Okabe, Daisuke Sano
    Water research 95 383 - 91 2016年05月15日 [査読有り][通常論文]
     
    We demonstrated the genotype-dependent removal of human norovirus particles with a microfiltration (MF) membrane in the presence of bacteria bearing histo-blood group antigens (HBGAs). Three genotypes (GII.3, GII.4, and GII.6) of norovirus-like particles (NoVLPs) were mixed with three bacterial strains (Enterobacter sp. SENG-6, Escherichia coli O86:K61:B7, and Staphylococcus epidermidis), respectively, and the mixture was filtered with an MF membrane having a nominal pore size of 0.45 μm. All NoVLP genotypes were rejected by the MF membrane in the presence of Enterobacter sp. SENG-6, which excreted HBGAs as extracellular polymeric substances (EPS). This MF membrane removal of NoVLPs was not significant when EPS was removed from cells of Enterobacter sp. SENG-6. GII.6 NoVLP was not rejected with the MF membrane in the presence of E. coli O86:K61:B7, but the removal of EPS of E. coli O86:K61:B7 increased the removal efficiency due to the interaction of NoVLPs with the exposed B-antigen in lipopolysaccharide (LPS) of E. coli O86:K61:B7. No MF membrane removal of all three genotypes was observed when S. epidermidis, an HBGA-negative strain, was mixed with NoVLPs. These results demonstrate that the location of HBGAs on bacterial cells is an important factor in determining the genotype-dependent removal efficiency of norovirus particles with the MF membrane. The presence of HBGAs in mixed liquor suspended solids from a membrane bioreactor (MBR) pilot plant was confirmed by immune-transmission electron microscopy, which implies that bacterial HBGAs can contribute to the genotype-dependent removal of human noroviruses with MBR using MF membrane.
  • 小林彩乃, 佐野大輔, 加藤毅, 伊藤寿宏, 宮村明帆, 三浦尚之, 石井聡, 岡部聡
    土木学会論文集 72 3 40 - 49 2016年03月 [査読有り][通常論文]
  • Tsuyoshi Kato, Ayano Kobayashi, Toshihiro Ito, Takayuki Miura, Satoshi Ishii, Satoshi Okabe, Daisuke Sano
    Journal of water and health 14 1 14 - 25 2016年02月 [査読有り][通常論文]
     
    A stochastic model for estimating the ratio between a fecal indicator and a pathogen based on left-censored data, which includes a substantially high number of non-detects, was constructed. River water samples were taken for 16 months at six points in a river watershed, and conventional fecal indicators (total coliforms and general Escherichia coli), genetic markers (Bacteroides spp.), and virulence genes (eaeA of enteropathogenic E. coli and ciaB of Campylobacter jejuni) were quantified. The quantification of general E. coli failed to predict the presence of the virulence gene from enteropathogenic E. coli, different from what happened with genetic markers (Total Bac and Human Bac). A Bayesian model that was adapted to left-censored data with a varying analytical quantification limit was applied to the quantitative data, and the posterior predictive distributions of the concentration ratio were predicted. When the sample size was 144, simulations conducted in this study suggested that 39 detects were enough to accurately estimate the distribution of the concentration ratio, when combined with a dataset with a positive rate higher than 99%. To evaluate the level of accuracy in the estimation, it is desirable to perform a simulation using an artificially generated left-censored dataset that has the identical number of non-detects as the actual data.
  • Rathnayake M L D Rathnayake, Mamoru Oshiki, Satoshi Ishii, Takahiro Segawa, Hisashi Satoh, Satoshi Okabe
    Bioresource technology 197 15 - 22 2015年12月 [査読有り][通常論文]
     
    The effects of dissolved oxygen (DO) and pH on nitrous oxide (N2O) production rates and pathways in autotrophic partial nitrification (PN) granules were investigated at the granular level. N2O was primarily produced by betaproteobacterial ammonia-oxidizing bacteria, mainly Nitrosomonas europaea, in the oxic surface layer (<200μm) of the autotrophic PN granules. N2O production increased with increasing bulk DO concentration owing to activation of the ammonia (i.e., hydroxylamine) oxidation in this layer. The highest N2O emissions were observed at pH 7.5, although the ammonia oxidation rate was unchanged between pH 6.5 and 8.5. Overall, the results of this study suggest that in situ analyses of PN granules are essential to gaining insight into N2O emission mechanisms in a granule.
  • Muhammad Ali, Satoshi Okabe
    Chemosphere 141 144 - 53 2015年12月 [査読有り][通常論文]
     
    Nitrogen removal from wastewater via anaerobic ammonium oxidation (anammox)-based process has been recognized as efficient, cost-effective and low energy alternative to the conventional nitrification and denitrification processes. To date, more than one hundred full-scale anammox plants have been installed and operated for treatment of NH4(+)-rich wastewater streams around the world, and the number is increasing rapidly. Since the discovery of anammox process, extensive researches have been done to develop various anammox-based technologies. However, there are still some challenges in practical application of anammox-based treatment process at full-scale, e.g., longer start-up period, limited application to mainstream municipal wastewater and poor effluent water quality. This paper aimed to summarize recent status of application of anammox process and researches on technological development for solving these remaining problems. In addition, an integrated system of anammox-based process and microbial fuel cell is proposed for sustainable and energy-positive wastewater treatment.
  • T. Ito, T. Kato, K. Takagishi, S. Okabe, D. Sano
    WATER SCIENCE AND TECHNOLOGY 72 10 1789 - 1795 2015年11月 [査読有り][通常論文]
     
    Left-censored datasets of virus density in wastewater samples make it difficult to evaluate the virus removal efficiency in wastewater treatment processes. In the present study, we modeled the probabilistic distribution of virus removal efficiency in a wastewater treatment process with a Bayesian approach, and investigated how many detect samples in influent and effluent are necessary for accurate estimation. One hundred left-censored data of virus density in wastewater (influent and effluent) were artificially generated based on assumed log-normal distributions and the posterior predictive distribution of virus density, and the log-ratio distribution were estimated. The estimation accuracy of distributions was quantified by Bhattacharyya coefficient. When it is assumed that the accurate estimation of posterior predictive distributions is possible when a 100% positive rate is obtained for 12 pairs of influent and effluent, 11 out of 144, 60 out of 324, and 201 out of 576 combinations of detect samples gave an accurate estimation at the significant level of 0.01 in a Kruskal-Wallis test when the total sample number was 12, 18, and 24, respectively. The combinations with the minimum number of detect samples were (12, 9), (16, 10), and (21, 8) when the total sample number was 12, 18, and 24, respectively.
  • Muhammad Ali, Mamoru Oshiki, Lashitha Rathnayake, Satoshi Ishii, Hisashi Satoh, Satoshi Okabe
    Water research 79 147 - 57 2015年08月01日 [査読有り][通常論文]
     
    Rapid start-up of anaerobic ammonium oxidation (anammox) process in up-flow column reactors was successfully achieved by immobilizing minimal quantity of biomass in polyvinyl alcohol (PVA)-sodium alginate (SA) gel beads. The changes in the reactor performance (i.e., nitrogen removal rate; NRR) were monitored with time. The results demonstrate that the reactor containing the immobilized biomass concentration of 0.33 g-VSS L(-1) achieved NRR of 10.8 kg-N m(-3) d(-1) after 35-day operation, whereas the reactor containing the granular biomass of 2.5 g-VSS L(-1) could achieve only NRR of 3.5 kg-N m(-3) d(-1). This indicates that the gel immobilization method requires much lower seeding biomass for start-up of anammox reactor. To explain the better performance of the immobilized biomass, the biological and physicochemical properties of the immobilized biomass were characterized and compared with the naturally aggregated granular biomass. Effective diffusion coefficient (De) in the immobilized biomass was directly determined by microelectrodes and found to be three times higher than one in the granular biomass. High anammox activity (i.e., NH4(+) and NO2(-) consumption rates) was evenly detected throughout the gel beads by microelectrodes due to faster and deeper substrate transport. In contrast, anammox activity was localized in the outer layers of the granular biomass, indicating that the inner biomass could not contribute to the nitrogen removal. This difference was in good agreement with the spatial distribution of microbes analysed by fluorescence in situ hybridization (FISH). Based on these results, PVA-SA gel immobilization is an efficient strategy to initiate anammox reactors with minimal quantity of anammox biomass.
  • Keitaro Yoshida, Yosuke Tashiro, Thithiwat May, Satoshi Okabe
    Water research 76 33 - 42 2015年06月01日 [査読有り][通常論文]
     
    In order to examine the interactions between physicochemical properties of specific extracellular polymeric substances (EPS) and membrane biofouling, we investigated the impacts of hydrophilic colanic acid, as a model extracellular polysaccharide component, on initial bacterial attachment to different microfiltration (MF) membranes and membrane biofouling by using Escherichia coli strains producing different amounts of colanic acid. In a newly designed microtiter plate assay, the bacterial attachment by an E. coli strain RcsF(+), which produces massive amounts of colanic acid, decreased only to a hydrophobic membrane because the colanic acid made cell surfaces more hydrophilic, resulting in low cell attachment to hydrophobic membranes. The bench-scale cross-flow filtration tests followed by filtration resistance measurement revealed that RcsF(+) caused severe irreversible membrane fouling (i.e., pore-clogging), whereas less extracellular polysaccharide-producing strains caused moderate but reversible fouling to all membranes used in this study. Further cross-flow filtration tests indicated that colanic acid liberated in the bulk phase could rapidly penetrate pre-accumulated biomass layers (i.e., biofilms) and then directly clogged membrane pores. These results indicate that colanic acid, a hydrophilic extracellular polysaccharide, and possible polysaccharides with similar characteristics with colanic acid are considered as a major cause of severe irreversible membrane fouling (i.e., pore-clogging) regardless of biofilm formation (dynamic membrane).
  • Muhammad Ali, Mamoru Oshiki, Takanori Awata, Kazuo Isobe, Zenichiro Kimura, Hiroaki Yoshikawa, Daisuke Hira, Tomonori Kindaichi, Hisashi Satoh, Takao Fujii, Satoshi Okabe
    Environmental microbiology 17 6 2172 - 89 2015年06月 [査読有り][通常論文]
     
    To date, six candidate genera of anaerobic ammonium-oxidizing (anammox) bacteria have been identified, and numerous studies have been conducted to understand their ecophysiology. In this study, we examined the physiological characteristics of an anammox bacterium in the genus 'Candidatus Jettenia'. Planctomycete KSU-1 was found to be a mesophilic (20-42.5°C) and neutrophilic (pH 6.5-8.5) bacterium with a maximum growth rate of 0.0020 h(-1) . Planctomycete KSU-1 cells showed typical physiological and structural features of anammox bacteria; i.e. (29) N2 gas production by coupling of (15) NH4 (+) and (14) NO2 (-) , accumulation of hydrazine with the consumption of hydroxylamine and the presence of anammoxosome. In addition, the cells were capable of respiratory ammonification with oxidation of acetate. Notably, the cells contained menaquinone-7 as a dominant respiratory quinone. Proteomic analysis was performed to examine underlying core metabolisms, and high expressions of hydrazine synthase, hydrazine dehydrogenase, hydroxylamine dehydrogenase, nitrite/nitrate oxidoreductase and carbon monoxide dehydrogenase/acetyl-CoA synthase were detected. These proteins require iron or copper as a metal cofactor, and both were dominant in planctomycete KSU-1 cells. On the basis of these experimental results, we proposed the name 'Ca. Jettenia caeni' sp. nov. for the bacterial clade of the planctomycete KSU-1.
  • Takayuki Miura, Satoshi Okabe, Yoshihito Nakahara, Daisuke Sano
    Water research 75 282 - 91 2015年05月15日 [査読有り][通常論文]
     
    In order to evaluate removal properties of human enteric viruses from wastewater by a membrane bioreactor (MBR), influent, anoxic and oxic mixed liquor, and membrane effluent samples were collected in a pilot-scale anoxic-oxic MBR process for 16 months, and concentrations of enteroviruses, norovirus GII, and sapoviruses were determined by real-time PCR using murine norovirus as a process control. Mixed liquor samples were separated into liquid and solid phases by centrifugation, and viruses in the bulk solution and those associated with mixed liquor suspended solids (MLSS) were quantified. Enteroviruses, norovirus GII, and sapoviruses were detected in the influent throughout the sampling period (geometrical mean, 4.0, 3.1, and 4.4 log copies/mL, respectively). Enterovirus concentrations in the solid phase of mixed liquor were generally lower than those in the liquid phase, and the mean log reduction value between influent and anoxic mixed liquor was 0.40 log units. In contrast, norovirus GII and sapovirus concentrations in the solid phase were equal to or higher than those in the liquid phase, and higher log reduction values (1.3 and 1.1 log units, respectively) were observed between influent and anoxic mixed liquor. This suggested that enteroviruses were less associated with MLSS than norovirus GII and sapoviruses, resulting in lower enterovirus removal in the activated sludge process. Enteroviruses and norovirus GII were detected in the MBR effluent but sapoviruses were not in any effluent samples. When MLSS concentration was reduced to 50-60% of a normal operation level, passages of enteroviruses and norovirus GII through a PVDF microfiltration membrane were observed. Since rejection of viruses by the membrane was not related to trans-membrane pressure which was monitored as a parameter of membrane fouling, the results indicated that adsorption to MLSS plays an important role in virus removal by an MBR, and removal properties vary by viruses reflecting different adsorptive behavior to MLSS. Our observations suggested that sapoviruses are more associated with MLSS and removed more efficiently than enteroviruses and norovirus GII.
  • Mamoru Oshiki, Kaori Shinyako-Hata, Hisashi Satoh, Satoshi Okabe
    Genome announcements 3 2 2015年04月16日 [査読有り][通常論文]
     
    A draft genome sequence of an anaerobic ammonium-oxidizing (anammox) bacterium, "Candidatus Brocadia sinica," was determined by pyrosequencing and by screening a fosmid library. A 4.07-Mb genome sequence comprising 3 contigs was assembled, in which 3,912 gene-coding regions, 47 tRNAs, and a single rrn operon were annotated.
  • Daisuke Sano, Takatomo Ohta, Arata Nakamura, Toyoko Nakagomi, Osamu Nakagomi, Satoshi Okabe
    Applied and environmental microbiology 81 8 2819 - 26 2015年04月 [査読有り][通常論文]
     
    The inability of molecular detection methods to distinguish disinfected virions from infectious ones has hampered the assessment of infectivity for enteric viruses caused by disinfection practices. In the present study, the reduction of infectivity of murine norovirus S7-PP3 and mengovirus vMC0, surrogates of human noroviruses and enteroviruses, respectively, caused by free-chlorine treatment was characterized culture independently by detecting carbonyl groups on viral capsid protein. The amount of carbonyls on viral capsid protein was evaluated by the proportion of biotinylated virions trapped by avidin-immobilized gel (percent adsorbed). This culture-independent approach demonstrated that the percent adsorbed was significantly correlated with the logarithm of the infectious titer of tested viruses. Taken together with the results of previous reports, the result obtained in this study indicates that the amount of carbonyls on viral capsid protein of four important families of waterborne pathogenic viruses, Astroviridae, Reoviridae, Caliciviridae, and Picornaviridae, is increased in proportion to the received oxidative stress of free chlorine. There was also a significant correlation between the percent adsorbed and the logarithm of the ratio of genome copy number to PFU, which enables estimation of the infectious titer of a subject virus by measuring values of the total genome copy number and the percent adsorbed. The proposed method is applicable when the validation of a 4-log reduction of viruses, a requirement in U.S. EPA guidelines for virus removal from water, is needed along with clear evidence of the oxidation of virus particles with chlorine-based disinfectants.
  • Akira Hafuka, Ryosuke Kando, Kohei Ohya, Koji Yamada, Satoshi Okabe, Hisashi Satoh
    BULLETIN OF THE CHEMICAL SOCIETY OF JAPAN 88 3 447 - 454 2015年03月 [査読有り][通常論文]
     
    In this paper, we investigated the effects of substitution at the 5-position of an asymmetric BODIPY cation sensor to tune its spectroscopic, photophysical, and cation-sensing properties. We introduced substituent groups with differing electron density at the 5-position of 3-[bis(pyridine-2-ylmethyl)amino]-BODIPY, which contains a cation recognition moiety at the 3-position of the BODIPY core, to develop four sensors which all exhibited distinctive ratiometric spectral changes in the presence of Cu2+. Aromatic substitution increased the Stokes shift. Substitution with the electron-withdrawing sulfonylphenyl group resulted in the highest fluorescence quantum yield, largest absorption coefficient, and largest spectral shift in the presence of Cu2+. The sulfonylphenyl-substituted sensor also exhibited excellent selectivity for Cu2+.
  • Oshiki, Mamoru, Shinyako-Hata, Kaori, Satoh, Hisashi, Okabe, Satoshi
    MICROBIOLOGY RESOURCE ANNOUNCEMENTS 3 2 2015年03月 [査読有り][通常論文]
     
    A draft genome sequence of an anaerobic ammonium-oxidizing (anammox) bacterium, "Candidatus Brocadia sinica," was determined by pyrosequencing and by screening a fosmid library. A 4.07-Mb genome sequence comprising 3 contigs was assembled, in which 3,912 gene-coding regions, 47 tRNAs, and a single rrn operon were annotated.
  • Satoshi Ishii, Gaku Kitamura, Takahiro Segawa, Ayano Kobayashi, Takayuki Miura, Daisuke Sano, Satoshi Okabe
    Applied and environmental microbiology 80 24 7505 - 11 2014年12月 [査読有り][通常論文]
     
    To secure food and water safety, quantitative information on multiple pathogens is important. In this study, we developed a microfluidic quantitative PCR (MFQPCR) system to simultaneously quantify 11 major human viral pathogens, including adenovirus, Aichi virus, astrovirus, enterovirus, human norovirus, rotavirus, sapovirus, and hepatitis A and E viruses. Murine norovirus and mengovirus were also quantified in our MFQPCR system as a sample processing control and an internal amplification control, respectively. River water contaminated with effluents from a wastewater treatment plant in Sapporo, Japan, was collected and used to validate our MFQPCR system for multiple viruses. High-throughput quantitative information was obtained with a quantification limit of 2 copies/μl of cDNA/DNA. Using this MFQPCR system, we could simultaneously quantify multiple viral pathogens in environmental water samples. The viral quantities obtained using MFQPCR were similar to those determined by conventional quantitative PCR. Thus, the MFQPCR system developed in this study can provide direct and quantitative information for viral pathogens, which is essential for risk assessments.
  • So Ishizaki, Itto Fujiki, Daisuke Sano, Satoshi Okabe
    Environmental science & technology 48 19 11204 - 10 2014年10月07日 [査読有り][通常論文]
     
    Alkalization on the cathode electrode limits the electrical power generation of air-cathode microbial fuel cells (MFCs), and thus external proton supply to the cathode electrode is essential to enhance the electrical power generation. In this study, the effects of external CO2 and water supplies to the cathode electrode on the electrical power generation were investigated, and then the relative contributions of CO2 and water supplies to the total proton consumption were experimentally evaluated. The CO2 supply decreased the cathode pH and consequently increased the power generation. Carbonate dissolution was the main proton source under ambient air conditions, which provides about 67% of total protons consumed for the cathode reaction. It is also critical to adequately control the water content on the cathode electrode of air-cathode MFCs because the carbonate dissolution was highly dependent on water content. On the basis of these experimental results, the power density was increased by 400% (143.0 ± 3.5 mW/m(2) to 575.0 ± 36.0 mW/m(2)) by supplying a humid gas containing 50% CO2 to the cathode chamber. This study demonstrates that the simultaneous CO2 and water supplies to the cathode electrode were effective to increase the electrical power generation of air-cathode MFCs for the first time.
  • 押木 守, 佐藤 久, 岡部 聡
    Journal of environmental biotechnology 14 1 21 - 29 環境バイオテクノロジー学会 2014年10月 [査読有り][通常論文]
  • Satoshi Ishii, Yanjun Song, Lashitha Rathnayake, Azzaya Tumendelger, Hisashi Satoh, Sakae Toyoda, Naohiro Yoshida, Satoshi Okabe
    Environmental microbiology 16 10 3168 - 80 2014年10月 [査読有り][通常論文]
     
    The identification of the key nitrous oxide (N2O) production pathways is important to establish a strategy to mitigate N2O emission. In this study, we combined real-time gas-monitoring analysis, (15)N stable isotope analysis, denitrification functional gene transcriptome analysis and microscale N2O concentration measurements to identify the main N2O producers in a partial nitrification (PN) aerobic granule reactor, which was fed with ammonium and acetate. Our results suggest that heterotrophic denitrification was the main contributor to N2O production in our PN aerobic granule reactor. The heterotrophic denitrifiers were probably related to Rhodocyclales bacteria, although different types of bacteria were active in the initial and latter stages of the PN reaction cycles, most likely in response to the presence of acetate. Hydroxylamine oxidation and nitrifier denitrification occurred, but their contribution to N2O emission was relatively small (20-30%) compared with heterotrophic denitrification. Our approach can be useful to quantitatively examine the relative contributions of the three pathways (hydroxylamine oxidation, nitrifier denitrification and heterotrophic denitrification) to N2O emission in mixed microbial populations.
  • Muhammad Ali, Mamoru Oshiki, Satoshi Okabe
    Water research 57 215 - 22 2014年06月15日 [査読有り][通常論文]
     
    It is still the biggest challenge to secure enough seeding biomass for rapid start-up of full-scale (anaerobic ammonium oxidation) anammox processes due to slow growth. Preservation of active anammox biomass could be one of the solutions. In this study, biomass of anammox bacterium, "Candidatus Brocadia sinica", immersed in various nutrient media were preserved at -80 °C, 4 °C and room temperature. After 45, 90 and 150 days of preservation, specific anammox activity (SAA) of the preserved anammox biomass was determined by measuring (29)N2 production rate and transcription levels of hzsA gene encoding hydrazine synthase alpha subunit. Storage in nutrient medium containing 3 mM of molybdate at room temperature with periodical (every 45 days) supply of NH4(+) and NO2(-) was proved to be the most effective storage technique for "Ca. Brocadia sinica" biomass. Using this preservation condition, 96, 92 and 65% of the initial SAA was sustained after 45, 90 and 150 days of storage, respectively. Transcription levels of hzsA gene in biomass correlated with the SAA (R(2) = 0.83), indicating it can be used as a genetic marker to evaluate the anammox activity of preserved biomass. Furthermore, the 90-day-stored biomass was successfully reactivated by immobilizing in polyvinyl alcohol (6%, w/v) and sodium alginate (2%, w/v) gel and then inoculated to up-flow column reactors. Total nitrogen removal rates rapidly increased to 7 kg-N m(-3) d(-1) within 35 days of operation. Based on these results, the room temperature preservation with molybdate addition is simple, cost-effective and feasible at a practical scale, which will accelerate the practical use of anammox process for wastewater treatment.
  • Zheng Ji, Xiaochang C. Wang, Limei Xu, Chongmiao Zhang, Naoyuki Funamizu, Satoshi Okabe, Daisuke Sano
    FOOD AND ENVIRONMENTAL VIROLOGY 6 2 99 - 109 2014年06月 [査読有り][通常論文]
     
    A polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) method was employed to estimate the contamination sources of human enteroviruses and understand how their dominant strains vary in a wastewater treatment and reclamation system consisting of sewage collection, wastewater treatment with membrane bioreactor and open lakes for reclaimed water storage and reuse. After PCR-DGGE using a selected primer set targeting enteroviruses, phylogenetic analysis of acquired enterovirus gene sequences was performed. Enteroviruses identified from the septic tank were much more diverse than those from grey water and kitchen wastewater. Several unique types of enterovirus different from those in wastewater samples were dominant in a biological wastewater treatment unit. Membrane filtration followed by chlorination was proved effective for physically eliminating enteroviruses; however, secondary contamination likely occurred as the reclaimed water was stored in artificial lakes. Enterovirus 71 (EV71), a hand-foot-and-mouth disease (HFMD) viral pathogen, was detected mainly from the artificial lakes, implying that wastewater effluent was not the contamination source of EV71 and that there were unidentified non-point sources of the contamination with the HFMD viral pathogen in the reclaimed water stored in the artificial lakes. The PCR-DGGE targeting enteroviruses provided robust evidence about viral contamination sources in the wastewater treatment and reclamation system.
  • Satoshi Ishii, Takamitsu Nakamura, Shuji Ozawa, Ayano Kobayashi, Daisuke Sano, Satoshi Okabe
    Environmental science & technology 48 9 4744 - 9 2014年05月06日 [査読有り][通常論文]
     
    Water quality monitoring and microbial risk assessment are important to ensure safe water for drinking, recreational, and agricultural purposes. In this study, we applied a microfluidic quantitative PCR (MFQPCR) approach to simultaneously quantify multiple waterborne pathogens in a natural freshwater lake in Hokkaido, Japan, from April to November, 2012. Tens of thousands of geese stopped over at this lake during their migration in spring and fall. Because lake water is used for irrigation of the surrounding agricultural area, we assessed infection risks through irrigation water usage based on pathogen concentrations directly measured by MFQPCR. We detected various pathogens in the lake water, particularly during the bird migration seasons, suggesting that migratory birds were the main source of the pathogens. However, neither counts of geese nor fecal indicator bacteria were good predictors of pathogen concentrations. On the basis of quantitative microbial risk assessment, concentrations of Campylobacter jejuni and Shigella spp. in water samples were above the concentrations that can potentially cause 10(-4) infections per person per year when water is used to grow fresh vegetables. These results suggest that direct and simultaneous multipathogen quantification can provide more reliable and comprehensive information for risk assessment than the current fecal indicator-based approach.
  • Akira Hafuka, Hiroaki Yoshikawa, Koji Yamada, Tsuyoshi Kato, Masahiro Takahashi, Satoshi Okabe, Hisashi Satoh
    Water research 54 12 - 20 2014年05月01日 [査読有り][通常論文]
     
    Fluorescence spectroscopy has great potential for on-site and real-time monitoring of pollutants in aquatic environments; however, its application to environmental aquatic samples has been extremely limited. In this study, a novel fluoroionophore based on a BODIPY-terpyridine conjugate was developed and applied to determine Zn concentrations in urban runoff. The fluoroionophore selectively bound to Zn(2+) in water, which led to an instant red-shift of the fluorescence peak of the fluoroionophore from 539 nm to 567 nm that could be seen by the naked eye. Zn concentrations could be quantified using the ratio of fluorescence intensities, and the detection limit was 9 μg/L, which is sufficiently low for environmental aquatic samples. To demonstrate applicability of the method to environmental samples, we measured Zn concentrations in urban runoff samples with a complex matrix (∼60 mg/L dissolved organic carbon and ∼20 mS/cm electrical conductivity). The total and dissolved fractions of Zn in the samples could be determined by fluorescence spectroscopy and its relative error was estimated to be less than 30% by inductively coupled plasma-atomic emission spectroscopy analysis. The proposed method is rapid and easy-to-use with simple pretreatment for Zn determination in environmental aquatic samples with complex matrices.
  • Zen-Ichiro Kimura, Kyung Mi Chung, Hiroaki Itoh, Akira Hiraishi, Satoshi Okabe
    International journal of systematic and evolutionary microbiology 64 Pt 4 1384 - 1388 2014年04月 [査読有り][通常論文]
     
    A Gram-stain-negative, non-spore-forming, rod-shaped bacterium, designated strain 1GB(T), was isolated from anodic biofilms of a glucose-fed microbial fuel cell. Strain 1GB(T) was facultatively anaerobic and chemo-organotrophic, having both a respiratory and a fermentative type of metabolism, and utilized a wide variety of sugars as carbon and energy sources. Cells grown aerobically contained Q-8 as the major quinone, but excreted Q-9 and a small amount of Q-10 when cultured with an electrode serving as the sole electron acceptor. The G+C content of the genomic DNA of 1GB(T) was 54.5 mol%. Multilocus sequence typing (MLST) analysis showed that strain 1GB(T) represented a distinct lineage within the genus Raoultella (98.5-99.4 % 16S rRNA gene sequence similarity and 94.0-96.5 % sequence similarity based on the three concatenated housekeeping genes gyrA, rpoB and parC. Strain 1GB(T) exhibited DNA-DNA hybridization relatedness of 7-43 % with type strains of all established species of the genus Raoultella. On the basis of these phenotypic, phylogenetic and genotypic data, the name Raoultella electrica sp. nov. is proposed for strain 1GB(T). The type strain is 1GB(T) ( = NBRC 109676(T) = KCTC 32430(T)).
  • Toshikazu Fukushima, Hiroe Hara-Yamamura, Makoto Urai, Ikuro Kasuga, Futoshi Kurisu, Taro Miyoshi, Katsuki Kimura, Yoshimasa Watanabe, Satoshi Okabe
    Water research 52 73 - 82 2014年04月01日 [査読有り][通常論文]
     
    Effects of chlorination on the toxicity of wastewater effluents treated by activated sludge (AS) and submerged membrane bioreactor (S-MBRB) systems to HepG2 human hepatoblastoma cells were investigated. In addition to the cytotoxicity and genotoxicity assays, the DNA microarray-based transcriptome analysis was performed to evaluate the change in types of biological impacts on HepG2 cells of the effluents by chlorination. Effluent organic matter (EfOM) and disinfection by-products (DBPs) were also characterized by using Fourier transform mass spectrometry (FT-MS). Although no significant induction of genotoxicity was observed by chlorination for both effluents, the chlorination elevated the cytotoxicity of AS effluent but reduced that of S-MBRB effluent. The FT-MS analyses revealed that more DBPs including nitrogenated DBPs (N-DBPs) were formed in the AS effluent than in the S-MBRB effluent by chlorination, supporting the increased cytotoxicity of AS effluent. The lower O/C ratio of S-MBRB EfOM suggests that a large number of organic molecules were detoxified by chlorination, which consequently decreased the cytotoxicity of S-MBRB effluent. Integration of all the results highlights that both cytotoxicity and biological impacts of chlorinated wastewater effluents were clearly dependent on the EfOM characteristics such as DBPs and O/C ratio, namely, on types of treatment systems.
  • 培養できないウイルスの感染性の評価法
    佐野大輔, 岡部聡
    感染と消毒 21 2 24 - 27 2014年 [査読無し][招待有り]
  • Tsuyoshi Kato, Takayuki Miura, Satoshi Okabe, Daisuke Sano
    FOOD AND ENVIRONMENTAL VIROLOGY 5 4 185 - 193 2013年12月 [査読有り][通常論文]
     
    Stochastic models are used to express pathogen density in environmental samples for performing microbial risk assessment with quantitative uncertainty. However, enteric virus density in water often falls below the quantification limit (non-detect) of the analytical methods employed, and it is always difficult to apply stochastic models to a dataset with a substantially high number of non-detects, i.e., left-censored data. We applied a Bayesian model that is able to model both the detected data (detects) and non-detects to simulated left-censored datasets of enteric virus density in wastewater. One hundred paired datasets were generated for each of the 39 combinations of a sample size and the number of detects, in which three sample sizes (12, 24, and 48) and the number of detects from 1 to 12, 24 and 48 were employed. The simulated observation data were assigned to one of two groups, i.e., detects and non-detects, by setting values on the limit of quantification to obtain the assumed number of detects for creating censored datasets. Then, the Bayesian model was applied to the censored datasets, and the estimated mean and standard deviation were compared to the true values by root mean square deviation. The difference between the true distribution and posterior predictive distribution was evaluated by Kullback-Leibler (KL) divergence, and it was found that the estimation accuracy was strongly affected by the number of detects. It is difficult to describe universal criteria to decide which level of accuracy is enough, but eight or more detects are required to accurately estimate the posterior predictive distributions when the sample size is 12, 24, or 48. The posterior predictive distribution of virus removal efficiency with a wastewater treatment unit process was obtained as the log ratio posterior distributions between the posterior predictive distributions of enteric viruses in untreated wastewater and treated wastewater. The KL divergence between the true distribution and posterior predictive distribution of virus removal efficiency also depends on the number of detects, and eight or more detects in a dataset of treated wastewater are required for its accurate estimation.
  • R. M. L. D. Rathnayake, Y. Song, A. Tumendelger, M. Oshiki, S. Ishii, H. Satoh, S. Toyoda, N. Yoshida, S. Okabe
    WATER RESEARCH 47 19 7078 - 7086 2013年12月 [査読有り][通常論文]
     
    Emission of nitrous oxide (N2O) during biological wastewater treatment is of growing concern since N2O is a major stratospheric ozone-depleting substance and an important greenhouse gas. The emission of N2O from a lab-scale granular sequencing batch reactor (SBR) for partial nitrification (PN) treating synthetic wastewater without organic carbon was therefore determined in this study, because PN process is known to produce more N2O than conventional nitrification processes. The average N2O emission rate from the SBR was 0.32 +/- 0.17 mgN L-1 h(-1), corresponding to the average emission of N2O of 0.8 +/- 0.4% of the incoming nitrogen load (1.5 +/- 0.8% of the converted NI-14"). Analysis of dynamic concentration profiles during one cycle of the SBR operation demonstrated that N2O concentration in off-gas was the highest just after starting aeration whereas N2O concentration in effluent was gradually increased in the initial 40 min of the aeration period and was decreased thereafter. Isotopomer analysis was conducted to identify the main N2O production pathway in the reactor during one cycle. The hydroxylamine (NH2OH) oxidation pathway accounted for 65% of the total N2O production in the initial phase during one cycle, whereas contribution of the NO reduction pathway to N2O production was comparable with that of the NH2OH oxidation pathway in the latter phase. In addition, spatial distributions of bacteria and their activities in single microbial granules taken from the reactor were determined with microsensors and by in situ hybridization. Partial nitrification occurred mainly in the oxic surface layer of the granules and ammonia-oxidizing bacteria were abundant in this layer. N2O production was also found mainly in the oxic surface layer. Based on these results, although N2O was produced mainly via NH2OH oxidation pathway in the autotrophic partial nitrification reactor, N2O production mechanisms were complex and could involve multiple N2O production pathways. (C) 2013 Elsevier Ltd. All rights reserved.
  • Takayuki Miura, Sylvain Parnaudeau, Marco Grodzki, Satoshi Okabe, Robert L. Atmar, Francoise S. Le Guyader
    APPLIED AND ENVIRONMENTAL MICROBIOLOGY 79 21 6585 - 6592 2013年11月 [査読有り][通常論文]
     
    Norovirus is the most common agent implicated in food-borne outbreaks and is frequently detected in environmental samples. These viruses are highly diverse, and three genogroups (genogroup I [GI], GII, and GIV) infect humans. Being noncultivable viruses, real-time reverse transcription-PCR (RT-PCR) is the only sensitive method available for their detection in food or environmental samples. Selection of consensus sequences for the design of sensitive assays has been challenging due to sequence diversity and has led to the development of specific real-time RT-PCR assays for each genogroup. Thus, sample screening can require several replicates for amplification of each genogroup (without considering positive and negative controls or standard curves). This study reports the development of a generic assay that detects all three human norovirus genogroups on a qualitative basis using a one-step real-time RT-PCR assay. The generic assay achieved good specificity and sensitivity for all three genogroups, detected separately or in combination. At variance with multiplex assays, the choice of the same fluorescent dye for all three probes specific to each genogroup allows the levels of fluorescence to be added and may increase assay sensitivity when multiple strains from different genogroups are present. When it was applied to sewage sample extracts, this generic assay successfully detected norovirus in all samples found to be positive by the genogroup-specific RT-PCRs. The generic assay also identified all norovirus-positive samples among 157 archived nucleic acid shellfish extracts, including samples contaminated by all three genogroups.
  • 羽深昭, 吉川弘晃, 大屋光平, 山田幸司, 高橋正宏, 岡部聡, 佐藤久
    土木学会論文集 50 3 275 - 280 2013年11月 [査読有り][通常論文]
  • Ayano Kobayashi, Daisuke Sano, Asami Taniuchi, Satoshi Ishii, Satoshi Okabe
    Applied microbiology and biotechnology 97 20 9165 - 73 2013年10月 [査読有り][通常論文]
     
    Quantitative PCR (qPCR) assays targeting the host-specific Bacteroides-Prevotella 16S rRNA genetic markers have been proposed as one of the promising approaches to identify the source of fecal contamination in environmental waters. One of the concerns of qPCR assays to environmental samples is the reliability of quantified values, since DNA extraction followed by qPCR assays are usually performed without appropriate sample process control (SPC) and internal amplification controls (IACs). To check the errors in sample processing and improve the reliability of qPCR results, it is essential to evaluate the DNA recovery efficiency and PCR amplification efficiency of the target genetic markers and correct the measurement results. In this study, we constructed a genetically-engineered Escherichia coli K12 strain (designated as strain MG1655 Δlac::kan) as sample process control and evaluated the applicability to environmental water samples. The recovery efficiency of the SPC strain MG1655 Δlac::kan was similar to that of Bacteroides fragilis JCM 11019, when DNA were extracted from water samples spiked with the two bacteria. Furthermore, the SPC was included in the qPCR assays with propidium monoazide (PMA) treatment, which can exclude the genetic markers from dead cells. No significant DNA loss was observed in the PMA treatment. The inclusion of both the SPC (strain MG1655 Δlac::kan) and IAC in qPCR assays with PMA treatment gave the assurance of reliable results of host-specific Bacteroides-Prevotella 16S rRNA genetic markers in environmental water samples.
  • Takayuki Miura, Daisuke Sano, Atsushi Suenaga, Takeshi Yoshimura, Miyu Fuzawa, Toyoko Nakagomi, Osamu Nakagomi, Satoshi Okabe
    Journal of virology 87 17 9441 - 51 2013年09月 [査読有り][通常論文]
     
    Histo-blood group antigens (HBGAs) have been suggested to be receptors or coreceptors for human noroviruses (HuNoVs) expressed on the intestinal epithelium. We isolated an enteric bacterium strain (SENG-6), closely related to Enterobacter cloacae, bearing HBGA-like substances from a fecal sample of a healthy individual by using a biopanning technique with anti-HBGA antibodies. The binding capacities of four genotypes of norovirus-like particles (NoVLPs) to Enterobacter sp. SENG-6 cells were confirmed by enzyme-linked immunosorbent assay (ELISA). Transmission electron microscopy demonstrated that NoVLPs bound mainly to extracellular polymeric substances (EPS) of Enterobacter sp. SENG-6, where the HBGA-like substances were localized. EPS that contained HBGA-like substances extracted from Enterobacter sp. SENG-6 was shown by enzyme-linked immunosorbent assay (ELISA) to be capable of binding to NoVLPs of a GI.1 wild-type strain (8fIIa) and a GII.6 strain that can recognize A antigen but not to an NoVLP GI.1 mutant strain (W375A) that loses the ability to bind to A antigen. Enzymatic cleavage of terminal N-acetyl-galactosamine residues in the bacterial EPS weakened bacterial EPS binding to the GI.1 wild-type strain (8fIIa). These results indicate that A-like substances in the bacterial EPS play a key role in binding to NoVLPs. Since the specific binding of HuNoVs to HBGA-positive enteric bacteria is likely to affect the transmission and infection processes of HuNoVs in their hosts and in the environment, further studies of human enteric bacteria and their binding capacity to HuNoVs will provide a new scientific platform for understanding interactions between two types of microbes that were previously regarded as biologically unrelated.
  • Ayano Kobayashi, Daisuke Sano, Jun Hatori, Satoshi Ishii, Satoshi Okabe
    Applied microbiology and biotechnology 97 16 7427 - 37 2013年08月 [査読有り][通常論文]
     
    Bacteroides-Prevotella group is one of the most promising targets for detecting fecal contamination in water environments, principally due to its host-specific distributions and high concentrations in feces of warm-blooded animals. We developed real-time PCR assays for quantifying chicken/duck-, chicken-, and duck-associated Bacteroides-Prevotella 16S rRNA genetic markers (Chicken/Duck-Bac, Chicken-Bac, and Duck-Bac). A reference collection of DNA extracts from 143 individual fecal samples and wastewater treatment plant influent was tested by the newly established markers. The quantification limits of Chicken/Duck-Bac, Chicken-Bac, and Duck-Bac markers in environmental water were 54, 57, and 12 copies/reaction, respectively. It was possible to detect possible fecal contaminations from wild ducks in environmental water with the constructed genetic marker assays, even though the density of total coliforms in the identical water samples was below the detection limit. Chicken/Duck-Bac marker was amplified from feces of wild duck and chicken with the positive ratio of 96 and 61 %, respectively, and no cross-reaction was observed for the other animal feces. Chicken-Bac marker was detected from 70 % of chicken feces, while detected from 39 % of cow feces, 8.3 % of pig feces, and 12 % of swan feces. Duck-Bac marker was detected from 85 % of wild duck feces and cross-reacted with 31 % of cow feces. These levels of detection specificity are common in avian-associated genetic markers previously proposed, which implies that there is a practical limitation in the independent application of avian-associated Bacteroides-Prevotella 16S rRNA genetic markers and a combination with other fecal contamination markers is preferable for detecting fecal contamination in water environments.
  • Zen-ichiro Kimura, Satoshi Okabe
    The ISME journal 7 8 1472 - 82 2013年08月 [査読有り][通常論文]
     
    Anodic microbial communities in acetate-fed microbial fuel cells (MFCs) were analyzed using stable-isotope probing of 16S rRNA genes followed by denaturing gradient gel electrophoresis. The results revealed that Geobacter sulfurreducens and Hydrogenophaga sp. predominated in the anodic biofilm. Although the predominance of Geobacter sp. as acetoclastic exoelectrogens in acetate-fed MFC systems has been often reported, the ecophysiological role of Hydrogenophaga sp. is unknown. Therefore, we isolated and characterized a bacterium closely related to Hydrogenophaga sp. (designated strain AR20). The newly isolated strain AR20 could use molecular hydrogen (H2), but not acetate, with carbon electrode as the electron acceptor, indicating that the strain AR20 was a hydrogenotrophic exoelectrogen. This evidence raises a hypothesis that acetate was oxidized by G. sulfurreducens in syntrophic cooperation with the strain AR20 as a hydrogen-consuming partner in the acetate-fed MFC. To prove this hypothesis, G. sulfurreducens strain PCA was cocultivated with the strain AR20 in the acetate-fed MFC without any dissolved electron acceptors. In the coculture MFC of G. sulfurreducens and strain AR20, current generation and acetate degradation were the highest, and the growth of strain AR20 was observed. No current generation, acetate degradation and cell growth occurred in the strain AR20 pure culture MFC. These results show for the first time that G. sulfurreducens can oxidize acetate in syntrophic cooperation with the isolated Hydrogenophaga sp. strain AR20, with electrode as the electron acceptor.
  • Takanori Awata, Mamoru Oshiki, Tomonori Kindaichi, Noriatsu Ozaki, Akiyoshi Ohashi, Satoshi Okabe
    APPLIED AND ENVIRONMENTAL MICROBIOLOGY 79 13 4145 - 4148 2013年07月 [査読有り][通常論文]
     
    The phylogenetic affiliation and physiological characteristics (e.g., K-s and maximum specific growth rate [mu(max)]) of an anaerobic ammonium oxidation (anammox) bacterium, "Candidatus Scalindua sp.," enriched from the marine sediment of Hiroshima Bay, Japan, were investigated. "Candidatus Scalindua sp." exhibits higher affinity for nitrite and a lower growth rate and yield than the known anammox species.
  • Yanjun Song, Satoshi Ishii, Lashitha Rathnayake, Tsukasa Ito, Hisashi Satoh, Satoshi Okabe
    Bioresource technology 139 285 - 91 2013年07月 [査読有り][通常論文]
     
    In this study, partial nitrifying (PN) aerobic granules were developed in a sequencing batch airlift reactor by controlling the airflow rate and NH4(+) loading rate. The PN reactor produced an effluent with a NO2(-)/NH4(+) ratio of approximately one and with an NH4(+) conversion rate of 1.22 kg N m(-3)day(-1). More than 95% of the total organic carbon was removed during the process. On the basis of clone library analysis and fluorescence in situ hybridization, ammonia-oxidizing bacteria (AOB) closely related to Nitrosomonas eutropha and putative heterotrophic denitrifiers were mainly present near the surface of the PN aerobic granules. Microelectrode measurements revealed that both NH4(+) and NO2(-) were consumed near the surface (<200 μm), whereas no nitrate (NO3(-)) accumulation was observed throughout the granules. These results indicate that PN by AOB and nitrite denitrification by heterotrophs, but not nitrite oxidation, simultaneously occurred near the surface of the PN aerobic granules.
  • M. Oshiki, S. Ishii, K. Yoshida, N. Fujii, M. Ishiguro, Hisashi Satoh, S. Okabe
    Applied and Environmental Microbiology 79 13 4087 - 4093 2013年07月 [査読有り][通常論文]
     
    We examined nitrate-dependent Fe2+ oxidation mediated by anaerobic ammonium oxidation (anammox) bacteria. Enrichment cultures of "Candidatus Brocadia sinica" anaerobically oxidized Fe2+ and reduced NO3- to nitrogen gasat rates of 3.7±0.2 and 1.3±0.1 (mean±standard deviation [SD]) nmol mg protein-1 min-1, respectively (37°C and pH 7.3). This nitrate reduction rate is an order of magnitude lower than the anammox activity of "Ca. Brocadia sinica" (10 to 75 nmol NH4+ mg protein-1 min-1). A 15N tracer experiment demonstrated that coupling of nitrate-dependent Fe2+ oxidation and the anammox reaction was responsible for producing nitrogen gas from NO3- by "Ca. Brocadia sinica." The activities of nitrate-dependent Fe2+ oxidation were dependent on temperature and pH, and thehighest activities were seen at temperatures of 30 to 45°C and pHs ranging from 5.9 to 9.8. The mean half-saturation constant for NO3 -±SD of "Ca. Brocadia sinica" was determined to be 51±21 μM. Nitrate-dependent Fe2+ oxidation was further demonstrated by another anammox bacterium, "Candidatus Scalindua sp.," whose rates of Fe2+ oxidation and NO3- reduction were 4.7±0.59 and 1.45±0.05 nmol mg protein-1 min-1, respectively (20°C and pH 7.3). Co-occurrence of nitrate-dependent Fe2+ oxidation and the anammox reaction decreased the molar ratios of consumed NO2 - to consumed NH4 + (δNO2-/δNH4 +) and produced NO3- to consumed NH4 + (δNO3 -/δNH4 +). These reactions are preferable to the application of anammox processes for wastewater treatment. © 2013, American Society for Microbiology.
  • Physiological characterization of an anaerobic ammonium-oxidizing bacterium enriched from coastal sediments,Hiroshima,Japan
    Takanori Awata, Mamoru Oshiki, Tomonori Kindaichi, Noriatsu Ozaki, Akiyoshi Ohashi, Satoshi Okabe
    The 2nd International Anammox Symposium pp.67  The 2nd International Anammox Symposium 2013年06月 [査読有り][通常論文]
     
    The effect of salinity on organic removal and ammonium oxidation in a down-flow hanging sponge reactor was investigated by conducting a long-term continuous experiment over a period of 800 days. The DHS reactor, constructed by connecting three identical units, was fed with artificial wastewater containing 500 mg-N/L of ammonium nitrogen and 1400 mg-COD/L of phenol. Salinity of the influent was controlled by the addition of 8.0 to 25 g-Cl-/L of NaCl. The DHS reactor was operated at a hydraulic retention time of 12 h in a temperature controlled room at 25 degrees C. No significant inhibition of organic removal and ammonium oxidation was observed at salinities of up to 20 g-Cl-/L, at which levels ammonium oxidation and COD removal both exceeded 90%, respectively. However, at a salinity of 25 g-Cl-/L, organic removal and ammonium oxidation were both severely inhibited. In addition, the ratio of effluent nitrite nitrogen to influent ammonium nitrogen increased from 3.4% at salinities of 8.0 g-Cl-/L to 33% at salinities of 20 g-Cl-/L.
  • Hiroe Hara-Yamamura, Koji Nakashima, Asiful Hoque, Taro Miyoshi, Katsuki Kimura, Yoshimasa Watanabe, Satoshi Okabe
    Environmental science & technology 47 10 5425 - 32 2013年05月21日 [査読有り][通常論文]
     
    DNA microarray-based transcriptome analysis with human hepatoma HepG2 cells was applied to evaluate the impacts of whole wastewater effluents from the membrane bioreactors (MBRs) and the activated sludge process (AS). In addition, the conventional bioassays (i.e., cytotoxicity tests and bioluminescence inhibition test), which were well-established for the evaluation of the overall effluent toxicity, were also performed for the same samples. Transcriptome analysis revealed that 2 to 926 genes, which were categorized to 0 to 225 biological processes, were differentially expressed after exposure to the effluents and the raw wastewater. Among the tested effluents, the effluent from a MBR operated at a relatively long solid retention time (i.e., 40 days) and small membrane pore size (i.e., 0.03 μm) showed the least impacts on the HepG2 even at the level comparable to tap water. The observed gene expression responses were in good agreement with the results of cytotoxicity tests, and provided additional molecular mechanistic information on adverse effects occurred in the sublethal region. Furthermore, the genes related to "lipid metabolism", "response to endogenous stimulus", and "response to inorganic substance" were selected as potential genetic markers, and their expression levels were quantified to evaluate the cellular impacts and treatability of wastewater effluents. Although the harmful impacts and innocuous impacts could not be distinguished at present, the results demonstrated that the DNA microarray-based transcriptome analysis with human HepG2 cells was a powerful tool to rapidly and comprehensively evaluate impacts of whole wastewater effluents.
  • Satoshi Ishii, Takahiro Segawa, Satoshi Okabe
    Applied and environmental microbiology 79 9 2891 - 8 2013年05月 [査読有り][通常論文]
     
    The direct quantification of multiple pathogens has been desired for diagnostic and public health purposes for a long time. In this study, we applied microfluidic quantitative PCR (qPCR) technology to the simultaneous detection and quantification of multiple food- and waterborne pathogens. In this system, multiple singleplex qPCR assays were run under identical detection conditions in nanoliter-volume chambers that are present in high densities on a chip. First, we developed 18 TaqMan qPCR assays that could be run in the same PCR conditions by using prevalidated TaqMan probes. Specific and sensitive quantification was achieved by using these qPCR assays. With the addition of two previously validated TaqMan qPCR assays, we used 20 qPCR assays targeting 10 enteric pathogens, a fecal indicator bacterium (general Escherichia coli), and a process control strain in the microfluidic qPCR system. We preamplified the template DNA to increase the sensitivity of the qPCR assays. Our results suggested that preamplification was effective for quantifying small amounts of the template DNA without any major impact on the sensitivity, efficiency, and quantitative performance of qPCR. This microfluidic qPCR system allowed us to detect and quantify multiple pathogens from fecal samples and environmental water samples spiked with pathogens at levels as low as 100 cells/liter. These results suggest that the routine monitoring of multiple pathogens in food and water samples is now technically feasible. This method may provide more reliable information for risk assessment than the current fecal contamination indicator approach.
  • W. M. K. R. T. W. Bandara, M. Ikeda, H. Satoh, M. Sasakawa, Y. Nakahara, M. Takahashi, S. Okabe
    WATER ENVIRONMENT RESEARCH 85 5 387 - 390 2013年05月 [査読有り][通常論文]
     
    The effectiveness of degasification using a degassing membrane to improve chemical oxygen demand (COD) removal efficiency was investigated using a bench-scale upflow anaerobic sludge blanket (UASB) reactor. Vacuum degasification was able to transfer dissolved gas in the bulk liquid of the UASB reactor inside the membrane. Such a process might provide thermodynamically favorable conditions for the degradation of organic compounds. The COD-removal efficiency improved from 83% during normal operation to 90% during the degassing operation.
  • Ayano Kobayashi, Daisuke Sano, Satoshi Okabe
    Water science and technology : a journal of the International Association on Water Pollution Research 67 4 838 - 45 2013年 [査読有り][通常論文]
     
    Genetic markers derived from Bacteroidales spp. have been proposed as promising indicators for fecal contamination in the water environment. However, little is known about the persistency of Bacteroidales spp. 16S rRNA genetic markers in the natural environment, which hampers the precise identification of fecal contamination sources. In this study, the persistency of human-specific Bacteroidales spp. genetic markers in river water was investigated during a 3-week agitation. The copy number of Bacteroidales spp. genetic marker was decreased with agitation time, and was very sensitive to water temperature. After the 3-week agitation, three clones of 18S rRNA gene related to Glaucoma scintillans, Spumella-like flagellate, and Colpidium campylum were acquired. The presence of predators that can prey on target bacteria could also be a critical factor affecting the quantified value of genetic markers. It is very important to take these factors, water temperature and the presence of predator, into account for predicting the fate of genetic markers to accurately identify fecal pollution sources.
  • Hisashi Satoh, Satoshi Okabe
    Microbes and environments 28 2 166 - 79 2013年 [査読有り][通常論文]
     
    The availability of benthic O2 plays a crucial role in benthic microbial communities and regulates many important biogeochemical processes. Burrowing activities of macrobenthos in the sediment significantly affect O2 distribution and its spatial and temporal dynamics in burrows, followed by alterations of sediment microbiology. Consequently, numerous research groups have investigated O2 dynamics in macrofaunal burrows. The introduction of powerful tools, such as microsensors and planar optodes, to sediment analysis has greatly enhanced our ability to measure O2 dynamics in burrows at high spatial and temporal resolution with minimal disturbance of the physical structure of the sediment. In this review, we summarize recent studies of O2-concentration measurements in burrows with O2 microsensors and O2 planar optodes. This manuscript mainly focuses on the fundamentals of O2 microsensors and O2 planar optodes, and their application in the direct measurement of the spatial and temporal dynamics of O2 concentrations in burrows, which have not previously been reviewed, and will be a useful supplement to recent literature reviews on O2 dynamics in macrofaunal burrows.
  • Kazuki Tojo, Daisuke Sano, Takayuki Miura, Toyoko Nakagomi, Osamu Nakagomi, Satoshi Okabe
    Water science and technology : a journal of the International Association on Water Pollution Research 67 10 2236 - 40 2013年 [査読有り][通常論文]
     
    This study developed a novel approach for evaluating the infectivity of enteric viruses without cell culture. Cumulative carbonyl groups on the viral capsid protein were labeled using biotin hydrazide, and the biotinylated virions were separated using a spin column filled with avidin-immobilized gel. Rotavirus was treated with free chlorine at an initial concentration of 0.3 mg/L for 3 min, and the log reduction in the infectious titer was 0.19 log (standard deviation, SD = 0.05). The log reduction of rotavirus treated with free chlorine at an initial concentration of 0.6 mg/L for 3 min was 2.6 log (SD = 0.37). No significant reductions in the amplicon copy numbers were observed in these free chlorine-treated samples. The recovery levels of intact virions in the first three fractions after biotin-avidin affinity chromatography were 76, 21, and 2.8%, while those of virions treated with free chlorine at an initial concentration of 0.3 mg/L for 3 min were 70, 23, and 5.6%. These results showed that the proposed approach could discriminate a 0.19 log infectivity-reduced population from an intact population, although no reduction in the amplicon copy number was observed. This novel method could be applied to noncultivatable enteric viruses such as human norovirus and sapovirus, and it could be very helpful for evaluating the viral inactivation efficiencies of intervention measures.
  • Akira Hafuka, Hiroki Taniyama, Sang-Hyun Son, Koji Yamada, Masahiro Takahashi, Satoshi Okabe, Hisashi Satoh
    BULLETIN OF THE CHEMICAL SOCIETY OF JAPAN 86 1 37 - 44 2013年01月 [査読有り][通常論文]
     
    Two novel asymmetric BODIPY fluoroionophores with dipicolylamine (BDP-DPA, dipicolylamine: bis(pyridyl-methyl)) and terpyridine (BDP-TPY) are described. These fluoroionophores display opposite wavelength responses on complexation with heavy metal ions. Furthermore, the fluorescence spectra vary depending on the ionic species. In particular, BDP-DPA shows a high affinity toward Cr3+ and upon complexation, the fluorescence spectrum blue-shifts from 591 to 566 nm. In contrast, BDP-TPY preferentially binds to Zn2+ and the fluorescence spectra red-shifts from 539 to 567 nm. BDP-TPY is the first example of asymmetric BODIPY with a pyridyl receptor at the 3 position showing red-shifted fluorescence by complexation with metal ions. The concentration of each metal ion was successfully determined by ratiometric measurement. The wavelength-responses characteristics of these fluoroionophores could be very useful in the development of novel ratiometric fluoroionophores for metal ions.
  • Zen-ichiro Kimura, Satoshi Okabe
    The Journal of general and applied microbiology 59 4 261 - 6 2013年 [査読有り][通常論文]
     
    A Gram-negative, non-spore-forming, rod-shaped bacterial strain, AR20(T), was isolated from anodic biofilms of an acetate-fed microbial fuel cell in Japan and subjected to a polyphasic taxonomic study. Strain AR20(T) grew optimally at pH 7.0-8.0 and 25°C. It contained Q-8 as the predominant ubiquinone and C16:0, summed feature 3 (C16:1ω7c and/or iso-C15:02OH), and C18:1ω7c as the major fatty acids. The DNA G+C content was 67.1 mol%. A neighbor-joining phylogenetic tree revealed that strain AR20(T) clustered with three type strains of the genus Hydrogenophaga (H. flava, H. bisanensis and H. pseudoflava). Strain AR20(T) exhibited 16S rRNA gene sequence similarity values of 95.8-97.7% to the type strains of the genus Hydrogenophaga. On the basis of phenotypic, chemotaxonomic and phylogenetic data, strain AR20(T) is considered a novel species of the genus Hydrogenophaga, for which the name Hydrogenophaga electricum sp. nov. is proposed. The type strain is AR20(T) (= KCTC 32195(T) = NBRC 109341(T)).
  • Mamoru Oshiki, Takanori Awata, Tomonori Kindaichi, Hisashi Satoh, Satoshi Okabe
    Microbes and environments 28 4 436 - 43 2013年 [査読有り][通常論文]
     
    Enrichment cultures of anaerobic ammonium oxidation (anammox) bacteria as planktonic cell suspensions are essential for studying their ecophysiology and biochemistry, while their cultivation is still laborious. The present study aimed to cultivate two phylogenetically distinct anammox bacteria, "Candidatus Brocadia sinica" and "Ca. Scalindua sp." in the form of planktonic cells using membrane bioreactors (MBRs). The MBRs were continuously operated for more than 250 d with nitrogen loading rates of 0.48-1.02 and 0.004-0.09 kgN m(-3) d(-1) for "Ca. Brocadia sinica" and "Ca. Scalindua sp.", respectively. Planktonic anammox bacterial cells were successfully enriched (>90%) in the MBRs, which was confirmed by fluorescence in-situ hybridization and 16S rRNA gene sequencing analysis. The decay rate and half-saturation constant for NO2(-) of "Ca. Brocadia sinica" were determined to be 0.0029-0.0081 d(-1) and 0.47 mgN L(-1), respectively, using enriched planktonic cells. The present study demonstrated that MBR enables the culture of planktonic anammox bacterial cells, which are suitable for studying their ecophysiology and biochemistry.
  • Wasala M K R T W Bandara, Tomonori Kindaichi, Hisashi Satoh, Manabu Sasakawa, Yoshihito Nakahara, Masahiro Takahashi, Satoshi Okabe
    Water research 46 17 5756 - 5764 2012年11月01日 [査読有り][通常論文]
     
    Anaerobic treatment is an attractive option for the biological treatment of municipal wastewater. In this study, municipal wastewater was anaerobically treated with a bench-scale upflow anaerobic sludge blanket (UASB) reactor at temperatures from 6 to 31 °C for 18 months to investigate total chemical oxygen demand (COD) removal efficiency, archaeal community structure, and dissolved methane (D-CH(4)) recovery efficiency. The COD removal efficiency was more than 50% in summer and below 40% in winter with no evolution of biogas. Analysis of the archaeal community structures of the granular sludge from the UASB using 16S rRNA gene-cloning indicated that after microorganisms had adapted to low temperatures, the archaeal community had a lower diversity and the relative abundance of acetoclastic methanogens decreased together with an increase in hydrogenotrophic methanogens. D-CH(4), which was detected in the UASB effluent throughout the operation, could be collected with a degassing membrane. The ratio of the collection to recovery rates was 60% in summer and 100% in winter. For anaerobic treatment of municipal wastewater at lower temperatures, hydrogenotrophic methanogens play an important role in COD removal and D-CH(4) can be collected to reduce greenhouse gas emissions and avoid wastage of energy resources.
  • Tsukasa Ito, Kazumi Yoshiguchi, Herto Dwi Ariesyady, Satoshi Okabe
    BIORESOURCE TECHNOLOGY 123 599 - 607 2012年11月 [査読有り][通常論文]
     
    We investigated the major phylogenetic groups and population size of glucose-, propionate-, and acetate-degrading bacteria in the glucose-degrading anaerobic digester sludge by stable-isotope probing analysis of 16S rRNA (RNA-SIP) with [C-13(6)]glucose followed by time course analysis of microautoradiography combined with fluorescent in situ hybridization (MAR-FISH) with [U-C-14]glucose. The results indicated that glucose was predominately degraded to CH4 and CO2 by glucose-degrading Propionibacterium and Olsenella that are belonging to the phylum Actinobacteria, propionate-degrading Smithella and Syntrophobacter, and acetate-degrading Methanosaeta and Synergistes group 4 in this anaerobic sludge. The population size of propionate degraders was the smallest among three trophic groups and the specific degradation rate of propionate was also low. The specific degradation rate of acetate was low even though their population size was comparable to the glucose degraders. These results could explain why the degradation of propionate and acetate was the rate-limiting step in methanogenic glucose degradation. (C) 2012 Elsevier Ltd. All rights reserved.
  • Zheng Ji, Xiaochang Wang, Chongmiao Zhang, Takayuki Miura, Daisuke Sano, Naoyuki Funamizu, Satoshi Okabe
    MICROBES AND ENVIRONMENTS 27 3 288 - 292 2012年09月 [査読有り][通常論文]
     
    Hand, foot and mouth disease (HFMD), caused by a group of enteric viruses such as Enterovirus 71 (EV71), Coxsackievirus A16 (CVA16) and Coxsackievirus A10 (CVA10), is heavily epidemic in East Asia. This research focused on investigating the occurrence of HFMD pathogens in domestic sewage and secondary effluent before disinfection in a wastewater treatment plant (WWTP) in Xi'an, the largest megacity in northwest China. In order to simultaneously detect all three HFMD pathogens, a semi-nested RT-PCR assay was constructed with a newly designed primer set targeting conservative gene regions from the 5' untranslated region (UTR) to VP2. As a result, 86% of raw sewage samples and 29% of the secondary effluent samples were positive for the HFMD viral gene, indicating that HFMD pathogens were highly prevalent in domestic wastewater and that they could also persist, even with lower probability, in the secondary effluent before disinfection. Of the three HFMD pathogens, CVA10 was positive in 48% of the total samples, while the occurrences of CVA16 and EV71 were 12% and 2%, respectively. It could thus be stated that CVA10 is the main HFMD pathogen prevailing in the study area, at least during the investigation period. High genetic diversity in the conservative gene region among the same serotype of the HFMD pathogen was identified by phylogenetic analysis, implying that this HFMD pathogen replicates frequently among the population excreting the domestic sewage.
  • Physiological characteristics of marine anammox bacteria enriched from sea sediments,Hiroshima,Japan.
    Takanori Awata, Tomonori Kindaichi, Noriatsu Ozaki, Akiyoshi Ohashi, Mamoru Oshiki, Satoshi Okabe
    14th International Symposium on Microbial Ecology(isme14) PS09.457A - PS09.457A 14th International Symposium on Microbial Ecology(isme14) 2012年08月19日 [査読有り][通常論文]
  • Yosuke Tashiro, Koji Kawata, Asami Taniuchi, Kenji Kakinuma, Thithiwat May, Satoshi Okabe
    Journal of bacteriology 194 5 1169 - 76 2012年03月 [査読有り][通常論文]
     
    Bacteria show remarkable adaptability under several stressful conditions by shifting themselves into a dormant state. Less is known, however, about the mechanism underlying the cell transition to dormancy. Here, we report that the transition to dormant states is mediated by one of the major toxin-antitoxin systems, RelEB, in a cell density-dependent manner in Escherichia coli K-12 MG1655. We constructed a strain, IKA121, which expresses the toxin RelE in the presence of rhamnose and lacks chromosomal relBE and rhaBAD. With this strain, we demonstrated that RelE-mediated dormancy is enhanced at high cell densities compared to that at low cell densities. The initiation of expression of the antitoxin RelB from a plasmid, pCA24N, reversed RelE-mediated dormancy in bacterial cultures. The activation of RelE increased the appearance of persister cells against β-lactams, quinolones, and aminoglycosides, and more persister cells appeared at high cell densities than at low cell densities. Further analysis indicated that amino acid starvation and an uncharacterized extracellular heat-labile substance promote RelE-mediated dormancy. This is a first report on the induction of RelE-mediated dormancy by high cell density. This work establishes a population-based dormancy mechanism to help explain E. coli survival in stressful environments.
  • Satoshi Okabe, Yoshiyuki Nakamura, Hisashi Satoh
    Microbes and environments 27 3 242 - 9 2012年 [査読有り][通常論文]
     
    The amount of oxygen released by Phragmites roots and the community structure and in situ activity of nitrifying bacteria in the root biofilms were analyzed by the combined use of 16S rRNA gene-cloning analysis, quantitative PCR (qPCR) assay and microelectrodes. Axial and radial O₂ microprofiles were obtained for individual roots of Phragmites in a horizontal flow reactor fed with artificial medium continuously. Axial O₂ profiles revealed that O₂ was released at a rate of 0.21 μmol O₂ cm⁻² (root surface area) h⁻¹ only in the apical region (up to ca. 40 mm from the root apex), where there was a high abundance (10⁷ to 10⁸ copies g⁻¹ biomass) of Nitrosomonas-like AOB and Nitrospira-like NOB. This abundance, however, sharply declined to the detection limit at positions more basal than 80 mm. Phylogenetic analysis based on 16S rRNA gene identified strains related to Nitrosomonas oligotropha and Nitrosomonas cryotolerans as the predominant AOB and strains related to Nitrospira marina and Nitrospira moscoviensis as the predominant NOB in the root biofilms. Based on radial O₂ microprofiles, the oxic region only extended about 0.5 mm into the surrounding sediment due to a high rate of O₂ consumption in the rhizosphere. The net NH₄⁺ and O₂ consumption rates in the apical region were higher than those determined at the oxic sediment surface in which the abundance of AOB and NOB was one order of magnitude lower than in the rhizosphere. These results clearly indicated that Phragmites root biofilms played an important role in nitrification in the waterlogged anoxic sediment.
  • Tsukasa Ito, Kazumi Yoshiguchi, Herto Dwi Ariesyady, Satoshi Okabe
    ISME JOURNAL 5 12 1844 - 1856 2011年12月 [査読有り][通常論文]
     
    Major acetate-utilizing bacterial and archaeal populations in methanogenic anaerobic digester sludge were identified and quantified by radioisotope-and stable-isotope-based functional analyses, microautoradiography-fluorescence in situ hybridization (MAR-FISH) and stable-isotope probing of 16S rRNA (RNA-SIP) that can directly link 16S rRNA phylogeny with in situ metabolic function. First, MAR-FISH with (14)C-acetate indicated the significant utilization of acetate by only two major groups, unidentified bacterial cells and Methanosaeta-like filamentous archaeal cells, in the digester sludge. To identify the acetate-utilizing unidentified bacteria, RNA-SIP was conducted with (13)C(6)-glucose and (13)C(3)-propionate as sole carbon source, which were followed by phylogenetic analysis of 16S rRNA. We found that bacteria belonging to Synergistes group 4 were commonly detected in both 16S rRNA clone libraries derived from the sludge incubated with (13)C-glucose and (13)C-propionate. To confirm that this bacterial group can utilize acetate, specific FISH probe targeting for Synergistes group 4 was newly designed and applied to the sludge incubated with (14)C-acetate for MAR-FISH. The MAR-FISH result showed that bacteria belonging to Synergistes group 4 significantly took up acetate and their active population size was comparable to that of Methanosaeta in this sludge. In addition, as bacteria belonging to Synergistes group 4 had high K(m) for acetate and maximum utilization rate, they are more competitive for acetate over Methanosaeta at high acetate concentrations (2.5-10mM). To our knowledge, it is the first time to report the acetate-utilizing activity of uncultured bacteria belonging to Synergistes group 4 and its competitive significance to acetoclastic methanogen, Methanosaeta.The ISME Journal (2011) 5, 1844-1856; doi:10.1038/ismej.2011.59; published online 12 May 2011
  • Thithiwat May, Satoshi Okabe
    Environmental microbiology 13 12 3149 - 62 2011年12月 [査読有り][通常論文]
     
    A variety of bacterial cell surface structures and quorum signalling molecules play a role in biofilm development in Escherichia coli. However, here we show that an engineered reduced-genome E. coli mutant that lacks 17.6% of the parental E. coli genome, including the genes involved in the synthesis of various cell surface structures, such as type 1 fimbriae, curli, exopolysaccharide polymers and the autoinducer-2 signalling molecule, is able to develop mature biofilms. Using temporal gene expression profiling, we investigated phenotypic changes in reduced-genome biofilms in relation with the genes encoding the synthesis of different amino acids that were differentially expressed during biofilm formation. We identified and characterized entB, marR, dosC, mcbR and yahK genes, as involved in biofilm formation by the reduced-genome E. coli. Of these, for a first time, we demonstrated that overproduction of entB and yahK, which encode an enterobactin for iron transport and a hypothetical oxidoreductase protein, respectively, promoted biofilm development and maturation. Our results indicate that specific types of genes contribute to phenotypic changes in reduced-genome E. coli biofilms. In addition, this work demonstrates that the functions of biofilm-specific genes could be analysed through experiments using the reduced-genome E. coli.
  • Satoshi Okabe, Mamoru Oshiki, Yoshitaka Takahashi, Hisashi Satoh
    Water research 45 19 6461 - 70 2011年12月01日 [査読有り][通常論文]
     
    Emission of nitrous oxide (N(2)O) during biological wastewater treatment is of growing concern. The emission of N(2)O from a lab-scale two-reactor partial nitrification (PN)-anammox reactor was therefore determined in this study. The average emission of N(2)O from the PN and anammox process was 4.0±1.5% (9.6±3.2% of the removed nitrogen) and 0.1±0.07% (0.14±0.09% of the removed nitrogen) of the incoming nitrogen load, respectively. Thus, a larger part (97.5%) of N(2)O was emitted from the PN reactor. The total amount of N(2)O emission from the PN reactor was correlated to nitrite (NO(2)(-)) concentration in the PN effluent rather than DO concentration. In addition, further studies were performed to indentify a key biological process that is responsible for N(2)O emission from the anammox process (i.e., granules). In order to characterize N(2)O emission from the anammox granules, the in situ N(2)O production rate was determined by using microelectrodes for the first time, which was related to the spatial organization of microbial community of the granule as determined by fluorescence in situ hybridization (FISH). Microelectrode measurement revealed that the active N(2)O production zone was located in the inner part of the anammox granule, whereas the active ammonium consumption zone was located above the N(2)O production zone. Anammox bacteria were present throughout the granule, whereas ammonium-oxidizing bacteria (AOB) were restricted to only the granule surface. In addition, addition of penicillin G that inhibits most of the heterotrophic denitrifiers and AOB completely inhibited N(2)O production in batch experiments. Based on these results obtained, denitrification by putative heterotrophic denitrifiers present in the inner part of the granule was considered the most probable cause of N(2)O emission from the anammox reactor (i.e., granules).
  • Takahiro Imai, Daisuke Sano, Takayuki Miura, Satoshi Okabe, Keishi Wada, Yoshifumi Masago, Tatsuo Omura
    BMC BIOTECHNOLOGY 11 123  2011年12月 [査読有り][通常論文]
     
    Background: Water contamination with human enteric viruses has posed human health risks all over the world. Reasonable and facile methodologies for recovering and quantifying infectious enteric viruses in environmental samples are needed to address the issues of waterborne viral infectious diseases. In this study, a bacterial protein that has a binding capability with several enteric viruses is discovered, and its binding characteristics were investigated for utilizing it as a viral adsorbent in virus recovery and detection technologies. Results: A gene of an enteric virus-binding protein (EVBP), derived from a monomer of a bacterial chaperon protein GroEL, was successfully acquired from a genomic DNA library of activated sludge microorganisms with nested PCR. Equilibrium dissociation constants between EVBP and norovirus-like particles (NoVLPs) of genotypes GI.7 and GII.4, estimated with quartz crystal microbalance method, were 240 and 210 nM, respectively. These values of equilibrium dissociation constant imply that the binding affinity between EVBP and NoVLPs is 1 to 3-log weaker than that in general antigen-antibody interactions, but about 2-log stronger than that in weak specific interactions of proteins with cations and organic polymers. The adsorptions of EVBP to norovirus, group A rotavirus and poliovirus type 1 were found to be significant in enzyme-linked immunosorbent assay. Meanwhile, the binding of native GroEL tetradecamer to viral particles was weaker than that of EVBP, presumably because of a steric hindrance. The small molecule of EVBP could have an advantage in the access to the surface of viral particles with rugged structure. Conclusions: EVBP that has a broad binding spectrum to enteric viruses was newly discovered. The broad binding characteristic of EVBP would allow us to utilize it as a novel adsorbent for detecting diverse enteric viruses in clinical and environmental samples.
  • Satoshi Okabe, Mamoru Oshiki, Yoshitaka Takahashi, Hisashi Satoh
    Bioresource technology 102 13 6801 - 7 2011年07月 [査読有り][通常論文]
     
    The partial nitrification reactor was successfully started up and operated stably for more than 250 days with a maximum nitrite production rate of 1.12 kg-Nm(-3)day(-1). The important factors for successful partial nitrification were high ammonium loading rate (>1.0 kg-Nm(-3)day(-1)) and relatively high pH (ca. 8.0), giving high free ammonia concentrations (>10mg NH(3)-NL(-1)). In addition, the air flow rate must be controlled at the ratio of air flow rate to ammonium loading rate below 0.1 (m(air)(3)day(-1))/(kg-Nm(-3)day(-1)). After the establishment of stable partial nitrification, the effluent NO(2)(-)-N/NH(4)(+)-N ratio and effluent NO(3)(-)-N concentration were 1.20 ± 0.33 and 1.2 ± 1.0mg-NL(-1), respectively, which was then fed into an granular-sludge anammox reactor. Consistent nitrogen removal was achieved for more than 250 days with a maximum nitrogen removal rate of 15.0 kg-TNm(-3)day(-1).
  • Mamoru Oshiki, Masaki Shimokawa, Naoki Fujii, Hisashi Satoh, Satoshi Okabe
    Microbiology (Reading, England) 157 Pt 6 1706 - 1713 2011年06月 [査読有り][通常論文]
     
    The present study investigated the phylogenetic affiliation and physiological characteristics of bacteria responsible for anaerobic ammonium oxidization (anammox); these bacteria were enriched in an anammox reactor with a nitrogen removal rate of 26.0 kg N m(-3) day(-1). The anammox bacteria were identified as representing 'Candidatus Brocadia sinica' on the basis of phylogenetic analysis of rRNA operon sequences. Physiological characteristics examined were growth rate, kinetics of ammonium oxidation and nitrite reduction, temperature, pH and inhibition of anammox. The maximum specific growth rate (μ(max)) was 0.0041 h(-1), corresponding to a doubling time of 7 days. The half-saturation constants (K(s)) for ammonium and nitrite of 'Ca. B. sinica' were 28±4 and 86±4 µM, respectively, higher than those of 'Candidatus Brocadia anammoxidans' and 'Candidatus Kuenenia stuttgartiensis'. The temperature and pH ranges of anammox activity were 25-45 °C and pH 6.5-8.8, respectively. Anammox activity was inhibited in the presence of nitrite (50 % inhibition at 16 mM), ethanol (91 % at 1 mM) and methanol (86 % at 1 mM). Anammox activities were 80 and 70 % of baseline in the presence of 20 mM phosphorus and 3 % salinity, respectively. The yield of biomass and dissolved organic carbon production in the culture supernatant were 0.062 and 0.005 mol C (mol NH (+)(-4))(-1), respectively. This study compared physiological differences between three anammox bacterial enrichment cultures to provide a better understanding of anammox niche specificity in natural and man-made ecosystems.
  • Wasala M K R T W Bandara, Hisashi Satoh, Manabu Sasakawa, Yoshihito Nakahara, Masahiro Takahashi, Satoshi Okabe
    Water research 45 11 3533 - 40 2011年05月 [査読有り][通常論文]
     
    In this study, we investigated the efficiency of dissolved methane (D-CH(4)) collection by degasification from the effluent of a bench-scale upflow anaerobic sludge blanket (UASB) reactor treating synthetic wastewater. A hollow-fiber degassing membrane module was used for degasification. This module was connected to the liquid outlet of the UASB reactor. After chemical oxygen demand (COD) removal efficiency of the UASB reactor became stable, D-CH(4) discharged from the UASB reactor was collected. Under 35 °C and a hydraulic retention time (HRT) of 10 h, average D-CH(4) concentration could be reduced from 63 mg COD L(-1) to 15 mg COD L(-1); this, in turn, resulted in an increase in total methane (CH(4)) recovery efficiency from 89% to 97%. Furthermore, we investigated the effects of temperature and HRT of the UASB reactor on degasification efficiency. Average D-CH(4) concentration was as high as 104 mg COD L(-1) at 15 °C because of the higher solubility of CH(4) gas in liquid; the average D-CH(4) concentration was reduced to 14 mg COD L(-1) by degasification. Accordingly, total CH(4) recovery efficiency increased from 71% to 97% at 15 °C as a result of degasification. Moreover, degasification tended to cause an increase in particulate COD removal efficiency. The UASB reactor was operated at the same COD loading rate, but different wastewater feed rates and HRTs. Although average D-CH(4) concentration in the UASB reactor was almost unchanged (ca. 70 mg COD L(-1)) regardless of the HRT value, the CH(4) discharge rate from the UASB reactor increased because of an increase in the wastewater feed rate. Because the D-CH(4) concentration could be reduced down to 12 ± 1 mg COD L(-1) by degasification at an HRT of 6.7 h, the CH(4) recovery rate was 1.5 times higher under degasification than under normal operation.
  • 押木守, 岡部聡
    生物の科学 遺伝 65 3 48 - 54 2011年05月01日 [査読無し][通常論文]
  • Thithiwat May, Kenji Tsuruta, Satoshi Okabe
    The ISME journal 5 4 771 - 5 2011年04月 [査読有り][通常論文]
     
    Escherichia coli carrying a natural conjugative F-plasmid generates F-pili mating pairs, which is important for early biofilm formation. In this study, we investigated the effect of male-specific filamentous single stranded DNA bacteriophage (f1) and RNA bacteriophage (MS2) on the formation of biofilms by E. coli carrying a natural conjugative F-plasmid. We showed that the early biofilm formation was completely inhibited by addition of the f1 phage, but not the MS2 phage. This suggests that the tip of F-pili is the specific attachment site for mating pairs formation and the side of F-pili has a non-obligatory role during biofilm formation. The inhibitory effect of the f1 phage was dependent on the time of addition during the biofilm formation. No inhibitory effect was observed when the f1 phages were added to the mature biofilms. This resistant mechanism of the mature biofilms could be attributed to the biofilm-specific phenotypes representing that the F-pili mating pairs were already formed and then the curli production commenced during the biofilm maturation. The pre-formed mating pairs seemed to resist the f1 phages. Altogether, our results indicate a close relationship between the presence of conjugative plasmid and male-specific bacteriophages within sessile biofilm communities, as well as the possibility of using the male-specific bacteriophages to control biofilm formation.
  • Takeshi Yamada, Kae Kikuchi, Toshihiro Yamauchi, Koji Shiraishi, Tsukasa Ito, Satoshi Okabe, Akira Hiraishi, Akiyoshi Ohashi, Hideki Harada, Yoichi Kamagata, Kazunori Nakamura, Yuji Sekiguchi
    APPLIED AND ENVIRONMENTAL MICROBIOLOGY 77 6 2081 - 2087 2011年03月 [査読有り][通常論文]
     
    A filamentous bulking of a methanogenic granular sludge caused by uncultured filamentous bacteria of the candidate phylum KSB3 in an upflow anaerobic sludge blanket (UASB) system has been reported. To characterize the physiological traits of the filaments, a polyphasic approach consisting of rRNA-based activity monitoring of the KSB3 filaments using the RNase H method and substrate uptake profiling using microautoradiography combined with fluorescence in situ hybridization (MAR-FISH) was conducted. On the basis of rRNA-based activity, the monitoring of a full-scale UASB reactor operated continuously revealed that KSB3 cells became active and predominant (up to 54% of the total 16S rRNA) in the sludge when the carbohydrate loading to the system increased. Batch experiments with a short incubation of the sludge with maltose, glucose, fructose, and maltotriose at relatively low concentrations (approximately 0.1 mM) in the presence of yeast extract also showed an increase in KSB3 rRNA levels under anaerobic conditions. MAR-FISH confirmed that the KSB3 cells took up radioisotopic carbons from [(14)C] maltose and [(14)C] glucose under the same incubation conditions in the batch experiments. These results suggest that one of the important ecophysiological characteristics of KSB3 cells in the sludge is carbohydrate degradation in wastewater and that high carbohydrate loadings may trigger an outbreak of KSB3 bacteria, causing sludge bulking in UASB systems.
  • Akira Hafuka, Kenji Sakaida, Hisashi Satoh, Masahiro Takahashi, Yoshimasa Watanabe, Satoshi Okabe
    Bioresource Technology 102 3 3551 - 3553 2011年02月 [査読有り][通常論文]
     
    We investigated the effects of different feeding regimens (1-pulse, stepwise, and continuous) of fermented food-waste liquid on polyhydroxybutyrate (PHB) production. The fermentation liquid was filtered with a membrane filter (pore size, 0.45. μm) to remove anaerobic microorganisms and solids and used as a carbon source for Cupriavidus necator. One-pulse feeding yielded the highest cell concentration of C. necator. However, the PHB concentration was higher in the stepwise- and continuous-feeding regimens. Therefore, the continuous-feeding regimen was used for continuous PHB production. PHB could be produced over 259 h (8 draw-fill cycles) with a maximal PHB content of 87%, but the PHB concentration and content decreased with an increase in the operation time. © 2010 Elsevier Ltd.
  • Daisuke Sano, Unai Perez-Sautu, Susana Guix, Rosa Maria Pinto, Takayuki Miura, Satoshi Okabe, Albert Bosch
    APPLIED AND ENVIRONMENTAL MICROBIOLOGY 77 3 1111 - 1114 2011年02月 [査読有り][通常論文]
     
    Human sapoviruses (SaVs) were quantified and characterized in an 18-month survey conducted along the Llobregat river catchment area in Spain. Sample types included freshwater, untreated and treated wastewater, and drinking water. All genogroups were recovered, and a seasonal distribution was observed. This is the first report of SaV quantification and genotyping in the environment outside Japan.
  • Akira Hafuka, Kenji Sakaida, Hisashi Satoh, Masahiro Takahashi, Yoshimasa Watanabe, Satoshi Okabe
    Bioresource technology 102 3 3551 - 3 2011年02月 [査読有り][通常論文]
     
    We investigated the effects of different feeding regimens (1-pulse, stepwise, and continuous) of fermented food-waste liquid on polyhydroxybutyrate (PHB) production. The fermentation liquid was filtered with a membrane filter (pore size, 0.45 μm) to remove anaerobic microorganisms and solids and used as a carbon source for Cupriavidus necator. One-pulse feeding yielded the highest cell concentration of C. necator. However, the PHB concentration was higher in the stepwise- and continuous-feeding regimens. Therefore, the continuous-feeding regimen was used for continuous PHB production. PHB could be produced over 259 h (8 draw-fill cycles) with a maximal PHB content of 87%, but the PHB concentration and content decreased with an increase in the operation time.
  • Sunja Cho, Naoki Fujii, Taeho Lee, Satoshi Okabe
    BIORESOURCE TECHNOLOGY 102 2 652 - 659 2011年01月 [査読有り][通常論文]
     
    Up-flow oxygen-controlled biofilm reactors equipped with a non-woven fabric support were used as a single reactor system for autotrophic nitrogen removal based on a combined partial nitrification and anaerobic ammonium oxidation (anammox) reaction. The up-flow biofilm reactors were initiated as either a partial nitrifying reactor or an anammox reactor, respectively, and simultaneous partial nitrification and anammox was established by careful control of the aeration rate. The combined partial nitrification and anammox reaction was successfully developed in both biofilm reactors without additional biomass inoculation. The reactor initiated as the anammox reactor gave a slightly higher and more stable mean nitrogen removal rate of 0.35 (+/- 0.19) kg-N m(-3) d(-1) than the reactor initiated as the partial nitrifying reactor (0.23 (+/- 0.16) kg-N m(-3) d(-1)). FISH analysis revealed that the biofilm in the reactor started as the anammox reactor were composed of anammox bacteria located in inner anoxic layers that were surrounded by surface aerobic AOB layers, whereas AOB and anammox bacteria were mixed without a distinguishable niche in the biofilm in the reactor started as the partial nitrifying reactor. However, it was difficult to efficiently maintain the stable partial nitrification owing to inefficient aeration in the reactor, which is a key to development of the combined partial nitrification and anammox reaction in a single biofilm reactor. (C) 2010 Elsevier Ltd. All rights reserved.
  • Kyungmi Chung, Itto Fujiki, Satoshi Okabe
    Bioresource technology 102 1 355 - 60 2011年01月 [査読有り][通常論文]
     
    A two-chamber MFC system was operated continuously for more than 500 days to evaluate effects of biofilm and chemical scale formation on the cathode electrode on power generation. A stable power density of 0.57 W/m(2) was attained after 200 days operation. However, the power density decreased drastically to 0.2 W/m(2) after the cathodic biofilm and chemical scale were removed. As the cathodic biofilm and chemical scale partially accumulated on the cathode, the power density gradually recovered with time. Microbial community structure of the cathodic biofilm was analyzed based on 16S rRNA clone libraries. The clones closely related to Xanthomonadaceae bacterium and Xanthomonas sp. in the Gammaproteobacteria subdivision were most frequently retrieved from the cathodic biofilm. Results of the SEM-EDX analysis revealed that the cation species (Na(+) and Ca(2+)) were main constituents of chemical scale, indicating that these cations diffused from the anode chamber through the Nafion membrane. However, an excess accumulation of the biofilm and chemical scale on the cathode exhibited adverse effects on the power generation due to a decrease in the active cathode surface area and an increase in diffusion resistance for oxygen. Thus, it is important to properly control the formation of chemical scale and biofilm on the cathode during long-term operation.
  • Satoshi Okabe, Hisashi Satoh, Tomonori Kindaichi
    Methods in enzymology 496 163 - 84 2011年 [査読有り][通常論文]
     
    This chapter aims to highlight the great potential of the combined use of microautoradiography (MAR) combined with fluorescent in situ hybridization (FISH) and microsensor technology in studies of complex multispecies nitrifying biofilms. The combination of FISH and microsensor technology is a powerful and reliable tool to link the spatial organization of microbial communities and their in situ function at community levels. MAR-FISH can be used to simultaneously examine the 16S rRNA-based phylogenetic identity and specific metabolic activity of cultivable or uncultivable microorganisms within complex microbial communities at a single-cell level. Information obtained at both resolution levels must be combined to draw a clear picture of a complex multispecies biofilm ecosystem. In addition, ecophysiological interactions among community members in complex multispecies biofilms can be investigated by tracing the fate of radiolabeled [(14)C] atom incorporated in nitrifying bacteria with MAR-FISH. The structure, function, and ecophysiological interactions among community members in complex multispecies nitrifying biofilms will be illustrated as an example of the combined use of MAR-FISH and microsensor technology.
  • Satoshi Okabe, Mamoru Oshiki, Yoichi Kamagata, Nobuyasu Yamaguchi, Masanori Toyofuku, Yutaka Yawata, Yosuke Tashiro, Nobuhiko Nomura, Hiroyuki Ohta, Moriya Ohkuma, Akira Hiraishi, Kiwamu Minamisawa
    MICROBES AND ENVIRONMENTS 25 4 230 - 240 2010年12月 [査読有り][通常論文]
     
    Ribosomal RNA (rRNA) sequence-based molecular techniques emerged in the late 1980s, which completely changed our general view of microbial life. Coincidentally, the Japanese Society of Microbial Ecology (JSME) was founded, and its official journal "Microbes and Environments (M&E)" was launched, in 1985. Thus, the past 25 years have been an exciting and fruitful period for M&E readers and microbiologists as demonstrated by the numerous excellent papers published in M&E. In this minireview, recent progress made in microbial ecology and related fields is summarized, with a special emphasis on 8 landmark areas; the cultivation of uncultured microbes, in situ methods for the assessment of microorganisms and their activities, biofilms, plant microbiology, chemolithotrophic bacteria in early volcanic environments, symbionts of animals and their ecology, wastewater treatment microbiology, and the biodegradation of hazardous organic compounds.
  • Thithiwat May, Akinobu Ito, Satoshi Okabe
    Molecular genetics and genomics : MGG 284 5 333 - 42 2010年11月 [査読有り][通常論文]
     
    The ecological role of horizontal gene transfer within biofilms has been recently investigated, and it has been reported that conjugation directly induces bacteria to form biofilms via expression of conjugative pili. In this report, we described the contribution of bacterial conjugation during biofilm formation by Escherichia coli harboring a natural IncF conjugative F plasmid (F(+)). We showed that cell-to-cell pili interactions through the homosexual mating-pair formation among F(+) × F(+) cells (namely, F(-) phenocopy phenomenon) promote E. coli biofilm formation at the early development stage. The presence of F(+) × F(+) population is the result from heterogeneity within biofilms leading to sessile bacteria that grow at different rates, in which the late-stationary phase cells acted as F(-) phenocopy cells. According to global transcriptional analysis, the biofilm lifestyle shared similar gene expression pattern with F(-) phenocopies. F(-) phenocopy cells expressed specific sets of chromosomal genes (e.g., genes for general stress response and two-component systems) that control the regulation regions of F transfer operon by blocking surface exclusion proteins and DNA transfer machineries. However, mating-pair proteins were stabilized and consequently promoted F(+) × F(+) pili assembly. Thus, F(-) phenocopy phenomenon is an effective adaptive behavior of bacterial cells during biofilm formation.
  • Shigeo Sasaki, Satoshi Okabe, Yuji Miyahara
    JOURNAL OF PHYSICAL CHEMISTRY B 114 46 14995 - 15002 2010年11月 [査読有り][通常論文]
     
    The solubility of N-isopropylacrylamide (NIPA) in water was found to discretely change at 25 degrees C. The highly concentrated NIPA solution separated into two solutions, the concentrations of which were higher than the solubility below 25 degrees C and lower than the solubility above 25 degrees C. The X-ray crystallographic analysis indicated that a NIPA crystal formed in the aqueous solution was H(2)O free. When the aqueous NIPA phase-separated solutions were cooled down to -90 degrees C at a rate faster than 35 degrees C/min, the glassy structure formed. On the other hand, the crystalline solid formation of NIPA and H(2)O were observed when the solutions were cooled to 50 degrees C at a slower rate than 3 degrees C/min. The DSC measurements of the phase-separated solutions revealed that the energy levels of NIPA were +15.2, +11.5, and -0.08 kJ/mol (regarding the crystalline solid state at 25 degrees C as the ground state) for the liquid state, the H(2)O-poor solution and the H(2)O-rich solution in the phase-separated state, respectively. The experimental results are explained in terms of the molecular assemblies of NIPA and H(2)O molecules in the solutions.
  • Sunja Cho, Yoshitaka Takahashi, Naoki Fujii, Yohei Yamada, Hisashi Satoh, Satoshi Okabe
    CHEMOSPHERE 78 9 1129 - 1135 2010年02月 [査読有り][通常論文]
     
    We investigated nitrogen removal performance and responsible microbial community in an anaerobic up-flow granular bed anammox reactor. The anammox reactor was operated more than 1 year. Biomass in the reactor formed granules after about 2 months of operation, and a sufficient amount of the granules was retained in the reactor with a metallic net to avoid biomass washout during the entire operation. The average diameter of the granules was 3.6 mm at day 310. After 8 months of operation, stable nitrogen removal (60%) was achieved at an average total inorganic nitrogen removal rate of 14 kg-N m(-3) d(-1). The phylogenetic analysis and fluorescence in situ hybridization results revealed that the anammox granules consisted of mono species of anammox bacteria, "Candidatus Brocadia-like species", affiliated with "Candidatus Brocadia anammoxidans" with 16S rRNA gene sequence similarity of 95.7%. The relative abundance of the anammox bacteria in the granules was more than 80% of the total bacteria stained with 4',6-diamidino-2-phenylindole. The anammox bacteria were present throughout the granules whereas the other bacterial groups. Chloroflexi-like filamentous bacteria and betaproteobacterial ammonia-oxidizing bacteria, were mainly present on the surface of the anammox granules and around the anammox bacterial clusters. The in situ anammox activity was detected mainly from near the surface of granules to the upper 800 pm of the granules with microsensors. The granular anammox biomass tolerated higher concentrations of nitrite (400 mg-N L(-1)) than did the homogenized biomass (200 mg-N L(-1)) probably due to substrate diffusion limitation. (C) 2009 Elsevier Ltd. All rights reserved.
  • Hisashi Satoh, Mitsunori Odagiri, Tsukasa Ito, Satoshi Okabe
    Water research 43 18 4729 - 39 2009年10月 [査読有り][通常論文]
     
    Microbially induced concrete corrosion (MICC) caused by sulfuric acid attack in sewer systems has been a serious problem for a long time. A better understanding of microbial community structures of sulfate-reducing bacteria (SRB) and sulfur-oxidizing bacteria (SOB) and their in situ activities is essential for the efficient control of MICC. In this study, the microbial community structures and the in situ hydrogen sulfide production and consumption rates within biofilms and corroded materials developed on mortar specimens placed in a corroded manhole was investigated by culture-independent 16S rRNA gene-based molecular techniques and microsensors for hydrogen sulfide, oxygen, pH and the oxidation-reduction potential. The dark-gray gel-like biofilm was developed in the bottom (from the bottom to 4 cm) and the middle (4-20 cm from the bottom of the manhole) parts of the mortar specimens. White filamentous biofilms covered the gel-like biofilm in the middle part. The mortar specimens placed in the upper part (30 cm above the bottom of the manhole) were corroded. The 16S rRNA gene-cloning analysis revealed that one clone retrieved from the bottom biofilm sample was related to an SRB, 12 clones and 6 clones retrieved from the middle biofilm and the corroded material samples, respectively, were related to SOB. In situ hybridization results showed that the SRB were detected throughout the bottom biofilm and filamentous SOB cells were mainly detected in the upper oxic layer of the middle biofilm. Microsensor measurements demonstrated that hydrogen sulfide was produced in and diffused out of the bottom biofilms. In contrast, in the middle biofilm the hydrogen sulfide produced in the deeper parts of the biofilm was oxidized in the upper filamentous biofilm. pH was around 3 in the corroded materials developed in the upper part of the mortar specimens. Therefore, it can be concluded that hydrogen sulfide provided from the bottom biofilms and the sludge settling tank was emitted to the sewer atmosphere, then oxidized to corrosive compounds in the upper and middle parts of the manhole, and only the upper part of the mortar specimens were corroded, because in the middle part of the manhole the generated corrosive compounds (e.g., sulfuric acid) was reduced in the deeper parts of the biofilm.
  • Akinobu Ito, Thithiwat May, Asami Taniuchi, Koji Kawata, Satoshi Okabe
    Biotechnology and bioengineering 103 5 975 - 83 2009年08月01日 [査読有り][通常論文]
     
    Although importance of the rpoS gene on biofilm formation by Escherichia coli has been suggested, there has not been any report showing where the rpoS is expressed during biofilm formation process. Since physiological state of the cells in the biofilms is considerably heterogeneous, the expression of the rpoS gene must be heterogeneous. In this study, in situ spatial expression of the rpoS gene during biofilm formation was investigated with an rpoS-gfp transcriptional fusion mutant strain. A ribosomal binding site and a gene encoding a green fluorescent protein were introduced into the downstream of the rpoS gene, which enabled us to observe the in situ spatial expression of the rpoS gene during biofilm formation processes without any disturbance of the rpoS expression. In the early stages of the biofilm formation process, the rpoS gene was expressed in the most of the cells. On the other hand, the rpoS expression was observed only at the outside of the biofilms during the late stages of the biofilm formation process. The in situ spatial expression of the rpoS gene in the biofilm was verified by quantifying the expression levels of the rpoS at the outside and the inside of the biofilms with the real time RT-PCR. In addition, global gene expression analysis was performed with DNA microarray to investigate physiological difference between the outside and the inside of the biofilms. This heterogeneous rpoS expression profile suggested that the cells at the outside of the biofilm need to express the rpoS to shift the physiological state to the stationary growth mode such as induction of various stress responses and suppression of the motility.
  • Koji Kawata, Masato Osawa, Satoshi Okabe
    Environmental science & technology 43 15 6046 - 51 2009年08月01日 [査読有り][通常論文]
     
    Although it has been reported that silver nanoparticles (Ag-NPs) have strong acute toxic effects to various cultured cells, the toxic effects at noncytotoxic doses are still unknown. We, therefore, evaluated in vitro toxicity of Ag-NPs at noncytotoxic doses in human hepatoma cell line, HepG2, based on cell viability assay, micronucleus test, and DNA microarray analysis. We also used polystyrene nanoparticles (PS-NPs) and silver carbonate (Ag2CO3) as test materials to compare the toxic effects with respect to different raw chemical composition and form of silver. The cell viability assay demonstrated that Ag-NPs accelerated cell proliferation at low doses (< 0.5 mg/L), which was supported by the DNA microarray analysis showing significant induction of genes associated with cell cycle progression. However, only Ag-NPs exposure exhibited a significant cytotoxicity at higher doses (> 1.0 mg/L) and induced abnormal cellular morphology, displaying cellular shrinkage and acquisition of an irregular shape. In addition, only Ag-NPs exposure increased the frequency of micronucleus formation up to 47.9 +/- 3.2% of binucleated cells, suggesting that Ag-NPs appear to cause much stronger damages to chromosome than PS-NPs and ionic Ag+. Cysteine, a strong ionic Ag+ ligand, only partially abolished the formation of micronuclei mediated by Ag-NPs and changed the gene expression, indicating that ionic Ag+ derived from Ag-NPs could not fully explain these biological actions. Based on these discussions, it is concluded that both "nanosized particle of Ag" as well as "ionic Ag+" contribute to the toxic effects of Ag-NPs.
  • Kyungmi Chung, Satoshi Okabe
    Applied microbiology and biotechnology 83 5 965 - 77 2009年07月 [査読有り][通常論文]
     
    A mediator-less three-stage two-chamber microbial fuel cell (MFC) system was developed and operated continuously for more than 1.5 years to evaluate continuous power generation while treating artificial wastewater containing glucose (10 mM) concurrently. A stable power density of 28 W/m(3) was attained with an anode hydraulic retention time of 4.5 h and phosphate buffer as the cathode electrolyte. An overall dissolved organic carbon removal ratio was about 85%, and coulombic efficiency was about 46% in this MFC system. We also analyzed the microbial community structure of anode biofilms in each MFC. Since the environment in each MFC was different due to passing on the products to the next MFC in series, the microbial community structure was different accordingly. The anode biofilm in the first MFC consisted mainly of bacteria belonging to the Gammaproteobacteria, identified as Aeromonas sp., while the Firmicutes dominated the anode biofilms in the second and third MFCs that were mainly fed with acetate. Cyclic voltammetric results supported the presence of a redox compound(s) associated with the anode biofilm matrix, rather than mobile (dissolved) forms, which could be responsible for the electron transfer to the anode. Scanning electron microscopy revealed that the anode biofilms were comprised of morphologically different cells that were firmly attached on the anode surface and interconnected each other with anchor-like filamentous appendages, which might support the results of cyclic voltammetry.
  • Feng Jiang, Derek Hoi-wai Leung, Shiyu Li, Guang-Hao Chen, Satoshi Okabe, Mark C. M. van Loosdrecht
    WATER RESEARCH 43 13 3187 - 3198 2009年07月 [査読有り][通常論文]
     
    This study developed a new sewer biofilm model to simulate the pollutant transformation and biofilm variation in sewers under aerobic, anoxic and anaerobic conditions. The biofilm model can describe the activities of heterotrophic, autotrophic, and sulfate-reducing bacteria (SRB) in the biofilm as well as the variations in biofilm thickness, the spatial profiles of SRB population and biofilm density. The model can describe dynamic biofilm growth, multiple biomass evolution and competitions among organic oxidation, denitrification, nitrification, sulfate reduction and sulfide oxidation in a heterogeneous biofilm growing in a sewer. The model has been extensively verified by three different approaches, including direct verification by measurement of the spatial concentration profiles of dissolved oxygen, nitrate, ammonia, and hydrogen sulfide in sewer biofilm. The spatial distribution profile of SRB in sewer biofilm was determined from the fluorescent in situ hybridization (FISH) images taken by a confocal laser scanning microscope (CLSM) and were predicted well by the model. (C) 2009 Elsevier Ltd. All rights reserved.
  • Akinobu Ito, Asami Taniuchi, Thithiwat May, Koji Kawata, Satoshi Okabe
    Applied and environmental microbiology 75 12 4093 - 100 2009年06月 [査読有り][通常論文]
     
    Biofilms are considered to be highly resistant to antimicrobial agents. Several mechanisms have been proposed to explain this high resistance of biofilms, including restricted penetration of antimicrobial agents into biofilms, slow growth owing to nutrient limitation, expression of genes involved in the general stress response, and emergence of a biofilm-specific phenotype. However, since combinations of these factors are involved in most biofilm studies, it is still difficult to fully understand the mechanisms of biofilm resistance to antibiotics. In this study, the antibiotic susceptibility of Escherichia coli cells in biofilms was investigated with exclusion of the effects of the restricted penetration of antimicrobial agents into biofilms and the slow growth owing to nutrient limitation. Three different antibiotics, ampicillin (100 microg/ml), kanamycin (25 microg/ml), and ofloxacin (10 microg/ml), were applied directly to cells in the deeper layers of mature biofilms that developed in flow cells after removal of the surface layers of the biofilms. The results of the antibiotic treatment analyses revealed that ofloxacin and kanamycin were effective against biofilm cells, whereas ampicillin did not kill the cells, resulting in regrowth of the biofilm after the ampicillin treatment was discontinued. LIVE/DEAD staining revealed that a small fraction of resistant cells emerged in the deeper layers of the mature biofilms and that these cells were still alive even after 24 h of ampicillin treatment. Furthermore, to determine which genes in the biofilm cells are induced, allowing increased resistance to ampicillin, global gene expression was analyzed at different stages of biofilm formation, the attachment, colony formation, and maturation stages. The results showed that significant changes in gene expression occurred during biofilm formation, which were partly induced by rpoS expression. Based on the experimental data, it is likely that the observed resistance of biofilms can be attributed to formation of ampicillin-resistant subpopulations in the deeper layers of mature biofilms but not in young colony biofilms and that the production and resistance of the subpopulations were aided by biofilm-specific phenotypes, like slow growth and induction of rpoS-mediated stress responses.
  • Koji Kawata, Ryuhei Shimazaki, Satoshi Okabe
    Environmental and molecular mutagenesis 50 1 46 - 59 2009年01月 [査読有り][通常論文]
     
    Carcinogenesis is an important chronic toxicity of metals and metalloids, although their mechanisms of action are still unclear. Comparison of gene expression patterns induced by carcinogenic metals, metalloids, and model carcinogens would give an insight into understanding of their carcinogenic mechanisms. In this study, we examined the gene expression alteration in human hepatoma cell line, HepG2, after exposing to two metals (cadmium and nickel), a metalloid (arsenic), and three model carcinogenic chemicals N-dimethylnitrosoamine (DMN), 12-O-tetradecanoylphorbol-13-acetate (TPA), and tetrachloroethylene (TCE) using DNA microarrays with 8,795 human genes. Of the genes altered by As, Cd, and Ni exposures, 31-55% were overlapped with those altered by three model carcinogenic chemical exposures in our experiments. In particular, the metals and metalloid shared certain characteristics with TPA and TCE in remarkable upregulations of the genes associated with progression of cell cycle, which might play a central role in As, Cd, and Ni carcinogenesis. This characteristic of gene expression alteration was partially counteracted by intracellular accumulation of vitamin C in As-exposed cells, whereas the number of cell-cycle associated genes was increased in Cd- and Ni-exposed cells. In our experimental conditions, ROS might have an accelerative effect on the cell proliferation mechanisms of As, but have an inhibitory effect on those of other two heavy metals. Furthermore, based on the results of Q-PCR, the oncogene PTTG1, which was upregulated by all carcinogenic chemical exposures in the array experiments, might be a useful biomarker for evaluation of the carcinogenesis of inorganic carcinogens.
  • Yutaka Yawata, Hideaki Maseda, Satoshi Okabe, Akinobu Ito, Isao Sawada, Hiroaki Kurashima, Hiroo Uchiyama, Nobuhiko Nomura
    MICROBES AND ENVIRONMENTS 24 4 338 - 342 2009年 [査読有り][通常論文]
     
    N-Octanoyl-L-homoserine lactone (C8-HSL) is an acyl-homoserine-lactone signal utilized in the quorum-sensing (QS) systems of Burkholderia cenocepacia and other bacterial species. Although also produced by Pseudomonas aeruginosa, its role in this species has not been elucidated. Here, we report that C8-HSL modulated antibiotic resistance and pyocyanin production in a P. aeruginosa efflux pump-deficient mutant. The rhl/las quorum-sensing system and qscR gene were both shown to be nonessential in the C8-HSL-induced changes in ofloxacin resistance, suggesting that P. aeruginosa possesses a distinct pathway to respond to C8-HSL.
  • Ichiro Yamada, Norio Yoshino, Akemi Tetsumura, Satoshi Okabe, Masayuki Enomoto, Kenichi Sugihara, Jiro Kumagai, Hitoshi Shibuya
    International Journal of Biomedical Imaging 2009 659836  2009年 [査読有り][通常論文]
     
    Purpose. To assess the accuracy of high-resolution MR imaging as a means of evaluating mural invasion and lymph node metastasis by colorectal carcinoma in surgical specimens. Materials and Methods. High-resolution T1-weighted and T2-weighted MR images were obtained in 92 surgical specimens containing 96 colorectal carcinomas. Results. T2-weighted MR images clearly depicted the normal colorectal wall as consisting of seven layers. In 90 (94%) of the 96 carcinomas the depth of mural invasion depicted by MR imaging correlated well with the histopathologic stage. Nodal signal intensity on T2-weighted images (93%) and nodal border contour (93%) were more accurate than nodal size (89%) as indicators of lymph node metastasis, and MR imaging provided the highest accuracy (94%-96%) when they were combined. Conclusion. High-resolution MR imaging is a very accurate method for evaluating both mural invasion and lymph node metastasis by colorectal carcinoma in surgical specimens. Copyright © 2009 Ichiro Yamada et al.
  • Thithiwat May, Satoshi Okabe
    Journal of bacteriology 190 22 7479 - 90 2008年11月 [査読有り][通常論文]
     
    It has been shown that Escherichia coli harboring the derepressed IncFI and IncFII conjugative F plasmids form complex mature biofilms by using their F-pilus connections, whereas a plasmid-free strain forms only patchy biofilms. Therefore, in this study we investigated the contribution of a natural IncF conjugative F plasmid to the formation of E. coli biofilms. Unlike the presence of a derepressed F plasmid, the presence of a natural IncF F plasmid promoted biofilm formation by generating the cell-to-cell mating F pili between pairs of F(+) cells (approximately two to four pili per cell) and by stimulating the formation of colanic acid and curli meshwork. Formation of colanic acid and curli was required after the initial deposition of F-pilus connections to generate a three-dimensional mushroom-type biofilm. In addition, we demonstrated that the conjugative factor of F plasmid, rather than a pilus synthesis function, was involved in curli production during biofilm formation, which promoted cell-surface interactions. Curli played an important role in the maturation process. Microarray experiments were performed to identify the genes involved in curli biosynthesis and regulation. The results suggested that a natural F plasmid was more likely an external activator that indirectly promoted curli production via bacterial regulatory systems (the EnvZ/OmpR two-component regulators and the RpoS and HN-S global regulators). These data provided new insights into the role of a natural F plasmid during the development of E. coli biofilms.
  • Yuki Miura, Satoshi Okabe
    Environmental science & technology 42 19 7380 - 6 2008年10月01日 [査読有り][通常論文]
     
    We, for the first time, quantitatively determined cell specific uptake activities of microbial products (bacterial cell detritus and extracellular polymeric substances, EPS) by the member of uncultured Chloroflexiby using a microautoradiography combined with fluorescence in situ hybridization (MAR-FISH) technique. For this MAR-FISH analysis, we prepared [14C]-labeled microbial products from biomass sludge obtained and bacterial strains (Pseudomonas sp. and Acinetobacter sp.) isolated from our pilot-scale membrane bioreactor (MBR) as tracer substrates, which probably represent the more realistic food source in the MBR. The quantitative MAR-FISH analyses clearly showed that most of the uncultured Chloroflexi could indeed uptake the bacterial detritus of the two isolated strains with rates of 1.7-3.5 x 10(-7) g-C microm-2-surface area h(-1) (corresponding to 1.2-1.7 mg-C-bacterial detritus L(-1) h(-1)) in the cultures, which were, however, about 2 orders of magnitude lower than the uptake rates of simple monosaccharides (mannose, arabinose, fucose, and galactose). Based on these results and their high abundance (more than 20% of total bacteria detected with EUB338-mixed probes), it could be estimated that the uncultured Chloroflexi contributes 38-51% of the total degradation of microbial products occurred in the MAR-FISH cultures.
  • Community structure and function of candidate division TM7 in a wastewater treatment plant.
    Tomonori Kindaichi, Hiroshi Kajihara, Takashi Yamamoto, Noriatsu Ozaki, Akiyoshi Ohashi, Satoshi Okabe
    12th International Symposium on Microbial Ecology (isme12) 2008年08月 [査読有り][通常論文]
  • Akinobu Ito, Thithiwat May, Koji Kawata, Satoshi Okabe
    Biotechnology and bioengineering 99 6 1462 - 71 2008年04月15日 [査読有り][通常論文]
     
    Presence of starved, stationary phase-like zones in biofilms seems to be an important factor for biofilm formation. In this study, roles of rpoS gene in the formation of Escherichia coli biofilms were investigated. E. coli MG1655 wild type (WT) and rpoS mutant (DeltarpoS) strains were used to compare biofilm formation capacity and global gene expression. Even though the DeltarpoS strain could attach and form microcolonies on glass surfaces, it could not establish mature biofilms. DNA microarray analysis revealed that WT biofilms (WBF) showed similar pattern of gene expression with WT planktonic stationary phase, whereas DeltarpoS biofilms (MBF) showed similar pattern of gene expression with WT planktonic exponential phase. Genes involved in energy metabolism (atpIBEFHAG, atpC, cydAB) and flagella synthesis (flgB, flgC, flhD, fliA, fliC, fliY) showed increased expression in the MBF, but not in the WBF. Moreover, genes involved in stress responses (blc, cspG, dinD poxB, wcaF, wcaI, and yfcF) showed increased expression in the WBF compared to the MBF. These results suggested that the rpoS gene contributed in maturation of E. coli biofilms through regulation of global gene expression including energy metabolism, motility, and stress responses.
  • Lucille Abad, Satoshi Okabe, Nlitsuhiro Shibayama, Hisaaki Kudo, Seiichi Saiki, Charito Aranilla, Lorna Relleve, Alumanda de la Rosa
    INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES 42 1 55 - 61 2008年01月 [査読有り][通常論文]
     
    The conformational associative properties Of kappa-, iota-, and lambda-carrageenan and agar with irradiation dose were studied by dynamic light scattering. The random scission of the carrageenans and agar by gamma irradiation resulted in the formation of polydispersed lower molecular weight fragments. At high doses, the system moves towards uniformity. Conformational change from coil to helix was observed in all carrageenans and agar at doses up to 100kGy. The con Fort national change in X-carrageenan may be due to the irregular and hybrid structure of this polysaccharide. Only agar and X-carrageenan still undergo conformational transition at a high dose of 200 kGy. Gelation is observed for kappa-, L-carrageenan up to a dose of 50 kGy while gelation is still observed at 100 kGy for agar. Increase in the hydrodynamic radius with decreasing temperatures for the non-irradiated carrageenans follows this order: lambda-carrageenan > kappa-carrageenan > iota-carrageenan. Slight increases in hydrodynamic radius were observed with irradiation. (c) 2007 Elsevier B.V. All rights reserved.
  • Yuki Miura, Yoshimasa Watanabe, Satoshi Okabe
    Environmental science & technology 41 22 7787 - 94 2007年11月15日 [査読有り][通常論文]
     
    We operated pilot-scale submerged membrane bioreactors (MBR) treating real municipal wastewater for over 3 months and observed an interesting phenomenon that carbohydrate concentrations in the MBRs rapidly increased, which consequently resulted in membrane fouling, when relative abundance of the member of uncultured Chloroflexi decreased from over 30% of total Bacteria to less than 10%. We, therefore, hypothesized that the uncultured Chloroflexi present in the MBRs could preferentially degrade carbohydrates and consequently prevent membrane fouling. To test this hypothesis, we investigated the phylogenetic identity, diversity, and in situ physiology (substrate utilization characteristics) of Chloroflexi residing in the MBR by using 16S rRNA gene sequencing analysis and microautoradiography combined with fluorescence in situ hybridization (MAR-FISH) technique. Most of the clones related to the phylum Chloroflexiwere affiliated with the Chloroflexi subphylum 1 containing only a few cultured representatives. The MAR-FISH revealed that the members of Chloroflexi were metabolically versatile and could preferentially utilize glucose and N-acetyl glucosamine (a main substantial constituent of the cell wall peptidoglycan) under oxic and anoxic conditions. The utilization of these compounds was low at low pH. These findings suggest that the members of Chloroflexi are ecologically significant in the MBR treating municipal wastewater and are responsible for degradation of SMP including carbohydrates and cellular materials, which consequently reduces membrane fouling potential.
  • Hisashi Satoh, Yuki Miura, Ikuo Tsushima, Satoshi Okabe
    Applied and environmental microbiology 73 22 7300 - 7 2007年11月 [査読有り][通常論文]
     
    The microbial community structure and spatial distribution of microorganisms and their in situ activities in anaerobic granules were investigated by 16S rRNA gene-based molecular techniques and microsensors for CH(4), H(2), pH, and the oxidation-reduction potential (ORP). The 16S rRNA gene-cloning analysis revealed that the clones related to the phyla Alphaproteobacteria (detection frequency, 51%), Firmicutes (20%), Chloroflexi (9%), and Betaproteobacteria (8%) dominated the bacterial clone library, and the predominant clones in the archaeal clone library were affiliated with Methanosaeta (73%). In situ hybridization with oligonucleotide probes at the phylum level revealed that these microorganisms were numerically abundant in the granule. A layered structure of microorganisms was found in the granule, where Chloroflexi and Betaproteobacteria were present in the outer shell of the granule, Firmicutes were found in the middle layer, and aceticlastic Archaea were restricted to the inner layer. Microsensor measurements for CH(4), H(2), pH, and ORP revealed that acid and H(2) production occurred in the upper part of the granule, below which H(2) consumption and CH(4) production were detected. Direct comparison of the in situ activity distribution with the spatial distribution of the microorganisms implied that Chloroflexi contributed to the degradation of complex organic compounds in the outermost layer, H(2) was produced mainly by Firmicutes in the middle layer, and Methanosaeta produced CH(4) in the inner layer. We determined the effective diffusion coefficient for H(2) in the anaerobic granules to be 2.66 x 10(-5) cm(2) s(-1), which was 57% in water.
  • Hisashi Satoh, Yuki Miura, Ikuo Tsushima, Satoshi Okabe
    Applied and Environmental Microbiology 73 22 7300 - 7307 2007年11月 [査読無し][通常論文]
     
    The microbial community structure and spatial distribution of microorganisms and their in situ activities in anaerobic granules were investigated by 16S rRNA gene-based molecular techniques and microsensors for CH4, H2, pH, and the oxidation-reduction potential (ORP). The 16S rRNA gene-cloning analysis revealed that the clones related to the phyla Alphaproteobacteria (detection frequency, 51%), Firmicutes (20%), Chloroflexi (9%), and Betaproteobacteria (8%) dominated the bacterial clone library, and the predominant clones in the archaeal clone library were affiliated with Methanosaeta (73%). In situ hybridization with oligonucleotide probes at the phylum level revealed that these microorganisms were numerically abundant in the granule. A layered structure of microorganisms was found in the granule, where Chloroflexi and Betaproteobacteria were present in the outer shell of the granule, Firmicutes were found in the middle layer, and aceticlastic Archaea were restricted to the inner layer. Microsensor measurements for CH4, H2, pH, and ORP revealed that acid and H2 production occurred in the upper part of the granule, below which H2 consumption and CH4 production were detected. Direct comparison of the in situ activity distribution with the spatial distribution of the microorganisms implied that Chloroflexi contributed to the degradation of complex organic compounds in the outermost layer, H2 was produced mainly by Firmicutes in the middle layer, and Methanosaeta produced CH4 in the inner layer. We determined the effective diffusion coefficient for H2 in the anaerobic granules to be 2.66 × 10-5 cm2 s-1, which was 57% in water. Copyright © 2007, American Society for Microbiology. All Rights Reserved.
  • Masahiko Annaka, Kanae Morishita, Satoshi Okabe
    JOURNAL OF PHYSICAL CHEMISTRY B 111 40 11700 - 11707 2007年10月 [査読有り][通常論文]
     
    We investigated the phase behavior and the microscopic structure of the colloidal complexes constituted from neutral/polyelectrolyte diblock copolymers and oppositely charged surfactant by dynamic light scattering (DLS) and small-angle neutron scattering (SANS). The neutral block is poly(N-isopropylacrylamide) (PNIPAM), and the polyelectrolyte block is negatively charged poly(acrylic acid) (PAA). In aqueous solution with neutral pH, PAA behaves as a weak polyelectrolyte, whereas PNIPAM is neutral and in good-solvent condition at ambient temperature, but in poor-solvent condition above similar to 32 degrees C. This block copolymer, PNIPAM-b-PAA with a narrow polydispersity, is studied in aqueous solution with an anionic surfactant, dodecyltrimethylammonium bromide (DTAB). For a low surfactant-to-polymer charge ratio Z lower than the critical value Z(C), the colloidal complexes are single DTAB micelles dressed by a few PNIPAM-b-PAA. Above Z(C), the colloidal complexes form a core-shell microstructure. The core of the complex consists of densely packed DTA(+) micelles, most likely connected between them by PAA blocks. The intermicellar distance of the DTA(+) micelles is similar to 39 angstrom, which is independent of the charge ratio Z as well as the temperature. The corona of the complex is constituted from the thermosensitive PNIPAM. At lower temperature the macroscopic phase separation is hindered by the swollen PNIPAM chains. Above the critical temperature T-C, the PNIPAM corona collapses leading to hydrophobic aggregates of the colloidal complexes.
  • Satoshi Okabe, Yoko Shimazu
    Applied microbiology and biotechnology 76 4 935 - 44 2007年09月 [査読有り][通常論文]
     
    Host-specific Bacteroides-Prevotella 16S rRNA genetic markers are promising alternative indicators for identifying the sources of fecal pollution because of their high abundance in the feces of warm-blooded animals and high host specificity. However, little is known about the persistence of these genetic markers in environments after being released into environmental waters. The persistence of feces-derived four different host-specific Bacteroides-Prevotella 16S rRNA genetic makers (total, human-, cow-, and pig-specific) in environmental waters was therefore investigated at different incubation temperatures (4, 10, 20, and 30 degrees C) and salinities (0, 10, 20, and 30 ppt) and then compared with the survival of conventional fecal-indicator organisms. The host-specific genetic markers were monitored by using real-time polymerase chain reaction (PCR) assays with specific primer sets. Each host-specific genetic marker showed similar responses in non-filtered river water and seawater: They persisted longer at lower temperatures and higher salinities. In addition, these markers did not increase in all conditions tested. Decay rates for indicator organisms were lower than those for host-specific genetic markers at temperature above 10 degrees C. Furthermore, we investigated whether the PCR-detectable 16S rRNA genetic markers reflect the presence of live target cells or dead target cells in environmental waters. The result revealed that the detection of the Bacteroides-Prevotella 16S rRNA genetic markers in environmental waters mainly reflected the presence of 'viable but non-culturable' Bacteroides-Prevotella cells. These findings indicate that seasonal and geographical variations in persistence of these host-specific Bacteroides-Prevotella 16S rRNA genetic markers must be considered when we use them as alternative fecal indicators in environmental waters.
  • Tomonori Kindaichi, Ikuo Tsushima, Yuji Ogasawara, Masaki Shimokawa, Noriatsu Ozaki, Hisashi Satoh, Satoshi Okabe
    APPLIED AND ENVIRONMENTAL MICROBIOLOGY 73 15 4931 - 4939 2007年08月 [査読有り][通常論文]
     
    We investigated autotrophic anaerobic ammonium-oxidizing (anammox) biofilms for their spatial organization, community composition, and in situ activities by using molecular biological techniques combined with microelectrodes. Results of phyllogenetic analysis and fluorescence in situ hybridization (FISH) revealed that "Brocadia"-like anammox bacteria that hybridized with the Amx820 probe dominated, with 60 to 92% of total bacteria in the upper part (< 1,000 mu m) of the biofilm, where high anammox activity was mainly detected with microelectrodes. The relative abundance of anammox bacteria decreased along the How direction of the reactor. FISH results also indicated that Nitrosomonas-, Nitrosospira-, and Nitrosococcus-like aerobic ammonia-oxidizing bacteria (AOB) and Nitrospira-like nitrite-oxidizing bacteria (NOB) coexisted with anammox bacteria and accounted for 13 to 21% of total bacteria in the biofilms. Microelectrode measurements at three points along the anammox reactor revealed that the NH4+, and NO2- consumption rates decreased from 0.68 and 0.64 mu mol cm(-2) h(-1) at P2 (the second port, 170 mm from the inlet port) to 0.30 and 0.35 mu mol cm(-2) h(-1) at P3 (the third port, 205 rum from the inlet port), respectively. No anammox activity was detected at P4 (the fourth port, 240 mm from the inlet port), even though sufficient amounts of NH4+ and NO2- and a high abundance of anammox bacteria were still present. This result could be explained by the inhibitory effect of organic compounds derived from biomass decay and/or produced by anammox and coexisting bacteria in the upper parts of the biofilm and in the upstream part of the reactor. The anammox activities in the biofilm determined by microelectrodes reflected the overall reactor performance. The several groups of aerobic AOB lineages, Nitrospira-like NOB, and Betaproteobacteria coexisting in the anammox biofilm might consume a trace amount of O-2, or organic compounds, which consequently established suitable microenvironments for anammox bacteria.
  • Olga Savichtcheva, Noriko Okayama, Satoshi Okabe
    Water research 41 16 3615 - 28 2007年08月 [査読有り][通常論文]
     
    Occurrence and prevalence of different bacterial enteric pathogens as well as their relationships with conventional (total and fecal coliforms) and alternative fecal indicators (host-specific Bacteroides 16S rRNA genetic markers) were investigated for various water samples taken from different sites with different degrees of fecal contamination. The results showed that a wide range of bacterial pathogens could be detected in both municipal wastewater treatment plant samples and in surface water samples. Logistic regression analysis revealed that total and human-specific Bacteroides 16S rRNA genetic markers showed significant predictive values for the presence of Escheriachia coli O-157, Salmonella, heat-labile enterotoxin (LT) of enterotoxigenic E. coli (ETEC), and heat-stable enterotoxin for human (STh) of ETEC. The probability of occurrence of these pathogenic bacteria became significantly high when the concentrations of human-specific and total Bacteroides 16S rRNA genetic markers exceeded 10(3) and 10(4) copies/100 mL. In contrast, Clostridium perfringens was detected at high frequency regardless of sampling sites and levels of Bacteroides 16S rRNA genetic markers. No genes related to Shigella spp., Staphylococcus aureus and Vibrio cholerae were detected in any samples analyzed in this study. Conventional indicator microorganisms had low levels of correlation with the presence of pathogens as compared with the alternative fecal indicators. These results suggested that real-time PCR-based measurement of alternative Bacteroides 16S rRNA genetic markers was a rapid and sensitive tool to identify host-specific fecal pollution and probably associated bacterial pathogens. However, since one fecal indicator might not represent the relative abundance of all pathogenic bacteria, viruses and protozoa, combined application of alternative indicators with conventional ones could provide more comprehensive pictures of fecal contamination, its source and association with pathogenic microorganisms.
  • Herto Dwi Ariesyady, Tsukasa Ito, Kazumi Yoshiguchi, Satoshi Okabe
    Applied microbiology and biotechnology 75 3 673 - 83 2007年06月 [査読有り][通常論文]
     
    The phylogenetic and functional diversity of syntrophic propionate-oxidizing bacteria (POB) present in an anaerobic digester was investigated by microautoradiography combined with fluorescent in situ hybridization (MAR-FISH) that can directly link 16S rRNA phylogeny with in situ metabolic function. The syntrophic POB community in the anaerobic digester sludge consisted of at least four phylogenetic groups: Syntrophobacter, uncultured short rod Smithella (Smithella sp. SR), uncultured long rod Smithella (Smithella sp. LR), and an unidentified group. The activities of these POB groups were dependent on the propionate concentrations. The uncultured Smithella sp. SR accounted for 52-62% of the total active POB under all the propionate concentrations tested (0.5-15 mM). In contrast, uncultured Smithella sp. LR was active only at lower propionate concentrations and became a dominant active POB at 0.5 mM of propionate. Syntrophobacter accounted for 16-31% of the total active POB above 2.5 mM propionate, whereas the active Syntrophobacter population became low (ca. 6%) at 0.5 mM of propionate. The anaerobic digester was operated in a fill and draw mode, resulting in periodical changes in propionate concentration ranging from 0 to 10 mM. These phylogenetically and functionally diverse, to some extent functionally redundant, active POB communities were dynamically responding to the periodical changes in propionate concentration.
  • Koji Kawata, Hiroyuki Yokoo, Ryuhei Shimazaki, Satoshi Okabe
    Environmental science & technology 41 10 3769 - 74 2007年05月15日 [査読有り][通常論文]
     
    Microarray technology is proving to be a useful tool to classify undefined environmental toxicants, to investigate underlying mechanisms of toxicity, and to identify candidate toxicant-specific genetic markers by examining global effects of putative toxicants on gene expression profiles. The aim of this study was to evaluate the toxicities of six heavy metals through the comparison with gene expression patterns induced by well-known chemicals. For this purpose, we first identified the genes altered specifically in HepG2 under the exposure of 2,3-dimethoxy-1,4-naphthoquinone (DMNQ), phenol, and N-nitrosodimethylamine (DMN), which were selected as the model chemicals, using DNA microarray. On the basis of the expression profiles of these genes, toxicities of six heavy metals, arsenic, cadmium, nickel, antimony, mercury, and chromium, were evaluated. The specific gene alteration and hierarchical clustering revealed that biological action of six heavy metals was clearly related to that of DMNQ which has been reported to be a reactive oxygen species (ROS) generating chemical and which induced the genes associated with cell proliferative responses. These results suggest that cell proliferative responses which are probably caused by ROS are a major apparent biological action of high-dose heavy metals, supporting the previous reports. Overall, a mechanism-based classification by DNA microarray would be an efficient method for evaluation of toxicities of environmental samples.
  • Ikuo Tsushima, Yuji Ogasawara, Tomonori Kindaichi, Hisashi Satoh, Satoshi Okabe
    Water research 41 8 1623 - 34 2007年04月 [査読有り][通常論文]
     
    To promptly establish anaerobic ammonium oxidation (anammox) reactors, appropriate seeding sludge with high abundance and activity of anammox bacteria was selected by quantifying 16S rRNA gene copy numbers of anammox bacteria by real-time quantitative PCR (RTQ-PCR) and batch culture experiments. The selected sludge was then inoculated into up-flow fixed-bed biofilm column reactors with nonwoven fabric sheets as biomass carrier and the reactor performances were monitored over 1 year. The anammox reaction was observed within 50 days and a total nitrogen removal rate of 26.0 kg-Nm(-3)day(-1) was obtained after 247 days. To our knowledge, such a high rate has never been reported before. Hydraulic retention time (HRT) and influent NH(4)(+) to NO(2)(-) molar ratio could be important determinant factors for efficient nitrogen removal in this study. The higher nitrogen removal rate was obtained at the shorter HRT and higher influent NH(4)(+)/NO(2)(-) molar ratio. After anammox reactors were fully developed, the community structure, spatial organization and in situ activity of the anammox biofilms were analyzed by the combined use of a full-cycle of 16S rRNA approach and microelectrodes. In situ hybridization results revealed that the probe Amx820-hybridized anaerobic anammox bacteria were distributed throughout the biofilm (accounting for more than 70% of total bacteria). They were associated with Nitrosomonas-like aerobic ammonia-oxidizing bacteria (AAOB) in the surface biofilm. The anammox bacteria present in this study were distantly related to the Candidatus Brocadia anammoxidans with the sequence similarity of 95%. Microelectrode measurements showed that a high in situ anammox activity (i.e., simultaneous consumption of NH(4)(+) and NO(2)(-)) of 4.45 g-N of (NH(4)(+)+NO(2)(-))m(-2)day(-1) was detected in the upper 800 microm of the biofilm, which was consistent with the spatial distribution of anammox bacteria.
  • Satoshi Okabe, Noriko Okayama, Olga Savichtcheva, Tsukasa Ito
    Applied microbiology and biotechnology 74 4 890 - 901 2007年03月 [査読有り][通常論文]
     
    Based on the comparative 16S rRNA gene sequence analysis of fecal DNAs, we identified one human-, three cow-, and two pig-specific Bacteroides-Prevotella 16S rRNA genetic markers, designed host-specific real-time polymerase chain reaction (real-time PCR) primer sets, and successfully developed real-time PCR assay to quantify the fecal contamination derived from human, cow, and pig in natural river samples. The specificity of each newly designed host-specific primer pair was evaluated on fecal DNAs extracted from these host feces. All three cow-specific and two pig-specific primer sets amplified only target fecal DNAs (in the orders of 9-11 log(10) copies per gram of wet feces), showing high host specificity. This real-time PCR assay was then applied to the river water samples with different fecal contamination sources and levels. It was confirmed that this assay could sufficiently discriminate and quantify human, cow, and pig fecal contamination. There was a moderate level of correlation between the Bacteroides-Prevotella group-specific 16S rRNA gene markers with fecal coliforms (r (2) = 0.49), whereas no significant correlation was found between the human-specific Bacteroides 16S rRNA gene with total and fecal coliforms. Using a simple filtration method, the minimum detection limits of this assay were in the range of 50-800 copies/100 ml. With a combined sample processing and analysis time of less than 8 h, this real-time PCR assay is useful for monitoring or identifying spatial and temporal distributions of host-specific fecal contaminations in natural water environments.
  • Satoshi Okabe, Mitsunori Odagiri, Tsukasa Ito, Hisashi Satoh
    Applied and environmental microbiology 73 3 971 - 80 2007年02月 [査読有り][通常論文]
     
    Microbially induced concrete corrosion (MICC) in sewer systems has been a serious problem for a long time. A better understanding of the succession of microbial community members responsible for the production of sulfuric acid is essential for the efficient control of MICC. In this study, the succession of sulfur-oxidizing bacteria (SOB) in the bacterial community on corroding concrete in a sewer system in situ was investigated over 1 year by culture-independent 16S rRNA gene-based molecular techniques. Results revealed that at least six phylotypes of SOB species were involved in the MICC process, and the predominant SOB species shifted in the following order: Thiothrix sp., Thiobacillus plumbophilus, Thiomonas intermedia, Halothiobacillus neapolitanus, Acidiphilium acidophilum, and Acidithiobacillus thiooxidans. A. thiooxidans, a hyperacidophilic SOB, was the most dominant (accounting for 70% of EUB338-mixed probe-hybridized cells) in the heavily corroded concrete after 1 year. This succession of SOB species could be dependent on the pH of the concrete surface as well as on trophic properties (e.g., autotrophic or mixotrophic) and on the ability of the SOB to utilize different sulfur compounds (e.g., H2S, S0, and S2O3(2-)). In addition, diverse heterotrophic bacterial species (e.g., halo-tolerant, neutrophilic, and acidophilic bacteria) were associated with these SOB. The microbial succession of these microorganisms was involved in the colonization of the concrete and the production of sulfuric acid. Furthermore, the vertical distribution of microbial community members revealed that A. thiooxidans was the most dominant throughout the heavily corroded concrete (gypsum) layer and that A. thiooxidans was most abundant at the highest surface (1.5-mm) layer and decreased logarithmically with depth because of oxygen and H2S transport limitations. This suggested that the production of sulfuric acid by A. thiooxidans occurred mainly on the concrete surface and the sulfuric acid produced penetrated through the corroded concrete layer and reacted with the sound concrete below.
  • Yuki Miura, Mirian Noriko Hiraiwa, Tsukasa Ito, Takanori Itonaga, Yoshimasa Watanabe, Satoshi Okabe
    Water research 41 3 627 - 37 2007年02月 [査読有り][通常論文]
     
    Bacterial community structures in pilot-scale conventional membrane bioreactors (CMBRs) and hybrid MBRs (HMBRs) which were combined with pre-coagulation/sedimentation were analyzed by polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) and fluorescence in situ hybridization (FISH) techniques. The results were compared with the community structure in a full-scale activated sludge (AS) process treating the same municipal wastewater. The Dice index (Cs) of similarity analysis of DGGE banding patterns demonstrated that the microbial community in AS was more similar to those in CMBR1 and CMBR2 than HMBR1 and HMBR2. This suggested that influent wastewater composition had a larger impact on bacterial community structures. Long-term community structure changes in the HMBRs and CMBRs were monitored and analyzed over 240 days by Non-metric multidimensional scaling (NMDS) analysis of DGGE banding patterns. The NMDS analysis revealed that both HMBRs and CMBRs had marked changes in community structures during the first about 100 days. Thereafter the perpetual fluctuations of bacterial community structures were observed in both HMBRs and CMBRs, even though the stable MBR performances (the performance was measured as membrane permeability and removal of dissolved organic carbon, DOC) were achieved. These results suggest that not only the stability, but also the adequate dynamics ("flexibility") of the bacterial community structure are important for the stable performance of the MBRs treating complex municipal wastewater.
  • Hisashi Satoh, Yoshiyuki Nakamura, Satoshi Okabe
    Applied and environmental microbiology 73 4 1341 - 8 2007年02月 [査読有り][通常論文]
     
    Influences of infaunal burrows constructed by the polychaete (Tylorrhynchus heterochaetus) on O(2) concentrations and community structures and abundances of ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB) in intertidal sediments were analyzed by the combined use of a 16S rRNA gene-based molecular approach and microelectrodes. The microelectrode measurements performed in an experimental system developed in an aquarium showed direct evidence of O(2) transport down to a depth of 350 mm of the sediment through a burrow. The 16S rRNA gene-cloning analysis revealed that the betaproteobacterial AOB communities in the sediment surface and the burrow walls were dominated by Nitrosomonas sp. strain Nm143-like sequences, and most of the clones in Nitrospira-like NOB clone libraries of the sediment surface and the burrow walls were related to the Nitrospira marina lineage. Furthermore, we investigated vertical distributions of AOB and NOB in the infaunal burrow walls and the bulk sediments by real-time quantitative PCR (Q-PCR) assay. The AOB and Nitrospira-like NOB-specific 16S rRNA gene copy numbers in the burrow walls were comparable with those in the sediment surfaces. These numbers in the burrow wall at a depth of 50 to 55 mm from the surface were, however, higher than those in the bulk sediment at the same depth. The microelectrode measurements showed higher NH(4)(+) consumption activity at the burrow wall than those at the surrounding sediment. This result was consistent with the results of microcosm experiments showing that the consumption rates of NH(4)(+) and total inorganic nitrogen increased with increasing infaunal density in the sediment. These results clearly demonstrated that the infaunal burrows stimulated O(2) transport into the sediment in which otherwise reducing conditions prevailed, resulting in development of high NH(4)(+) consumption capacity. Consequently, the infaunal burrow became an important site for NH(4)(+) consumption in the intertidal sediment.
  • Ikuo Tsushima, Tomonori Kindaichi, Satoshi Okabe
    Water Research 41 4 785 - 794 2007年02月 [査読有り][通常論文]
     
    The anaerobic ammonium-oxidizing (ANAMMOX) bacteria were enriched from a rotating disk reactor (RDR) biofilm in semi-batch cultures. Based on fluorescence in situ hybridization (FISH) analysis, this enrichment led to a relative population size of 36% ANAMMOX bacteria. Phylogenetic analysis revealed that all the detected clones were related to the previously reported ANAMMOX bacteria, Candidatus Brocadia anammoxidans (AF375994), with 92% sequence similarity. Furthermore, we successfully developed a real-time polymerase chain reaction (PCR) assay to quantify populations of ANAMMOX bacteria in the enrichment cultures. For this real-time PCR assay, PCR primer sets targeting 16S ribosomal RNA genes of ANAMMOX bacteria were designed and used. The quantification range of this assay was 6 orders of magnitude, from 8.9×101 to 8.9×106 copies per PCR, corresponding to the detection limit of 3.6×103 target copies mL-1. A significant correlation was found between the increase in copy numbers of 16S rRNA gene of ANAMMOX bacteria and the increase in nitrogen removal rates in the enrichment cultures. Quantifying ANAMMOX bacterial populations in the enrichment culture made it possible to estimate the doubling time of the enriched ANAMMOX bacteria to be 3.6 to 5.4 days. The real-time PCR assay gave comparable population sizes in the enrichment cultures with the FISH results. These results suggest that the real-time PCR assay developed in this study is useful and reliable for quantifying the populations of ANAMMOX bacteria in environmental and engineering samples. © 2006 Elsevier Ltd. All rights reserved.
  • Ikuo Tsushima, Tomonori Kindaichi, Satoshi Okabe
    Water research 41 4 785 - 94 2007年02月 [査読有り][通常論文]
     
    The anaerobic ammonium-oxidizing (ANAMMOX) bacteria were enriched from a rotating disk reactor (RDR) biofilm in semi-batch cultures. Based on fluorescence in situ hybridization (FISH) analysis, this enrichment led to a relative population size of 36% ANAMMOX bacteria. Phylogenetic analysis revealed that all the detected clones were related to the previously reported ANAMMOX bacteria, Candidatus Brocadia anammoxidans (AF375994), with 92% sequence similarity. Furthermore, we successfully developed a real-time polymerase chain reaction (PCR) assay to quantify populations of ANAMMOX bacteria in the enrichment cultures. For this real-time PCR assay, PCR primer sets targeting 16S ribosomal RNA genes of ANAMMOX bacteria were designed and used. The quantification range of this assay was 6 orders of magnitude, from 8.9x10(1) to 8.9x10(6) copies per PCR, corresponding to the detection limit of 3.6x10(3) target copies mL(-1). A significant correlation was found between the increase in copy numbers of 16S rRNA gene of ANAMMOX bacteria and the increase in nitrogen removal rates in the enrichment cultures. Quantifying ANAMMOX bacterial populations in the enrichment culture made it possible to estimate the doubling time of the enriched ANAMMOX bacteria to be 3.6 to 5.4 days. The real-time PCR assay gave comparable population sizes in the enrichment cultures with the FISH results. These results suggest that the real-time PCR assay developed in this study is useful and reliable for quantifying the populations of ANAMMOX bacteria in environmental and engineering samples.
  • Hisashi Satoh, Yoshiyuki Nakamura, Satoshi Okabe
    Applied and Environmental Microbiology 73 4 1341 - 1348 2007年02月 [査読無し][通常論文]
     
    Influences of infaunal burrows constructed by the polychaete (Tylorrhynchus heterochaetus) on O2 concentrations and community structures and abundances of ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB) in intertidal sediments were analyzed by the combined use of a 16S rRNA gene-based molecular approach and microelectrodes. The microelectrode measurements performed in an experimental system developed in an aquarium showed direct evidence of O2 transport down to a depth of 350 mm of the sediment through a burrow. The 16S rRNA gene-cloning analysis revealed that the betaproteobacterial AOB communities in the sediment surface and the burrow walls were dominated by Nitrosomonas sp. strain Nm143-like sequences, and most of the clones in Nitrospira-like NOB clone libraries of the sediment surface and the burrow walls were related to the Nitrospira marina lineage. Furthermore, we investigated vertical distributions of AOB and NOB in the infaunal burrow walls and the bulk sediments by real-time quantitative PCR (Q-PCR) assay. The AOB and Nitrospira-like NOB-specific 16S rRNA gene copy numbers in the burrow walls were comparable with those in the sediment surfaces. These numbers in the burrow wall at a depth of 50 to 55 mm from the surface were, however, higher than those in the bulk sediment at the same depth. The microelectrode measurements showed higher NH4+ consumption activity at the burrow wall than those at the surrounding sediment. This result was consistent with the results of microcosm experiments showing that the consumption rates of NH 4+ and total inorganic nitrogen increased with increasing infaunal density in the sediment. These results clearly demonstrated that the infaunal burrows stimulated O2 transport into the sediment in which otherwise reducing conditions prevailed, resulting in development of high NH4+ consumption capacity. Consequently, the infaunal burrow became an important site for NH4+ consumption in the intertidal sediment. Copyright © 2007, American Society for Microbiology. All Rights Reserved.
  • Yuki Miura, Yoshimasa Watanabe, Satoshi Okabe
    Environmental science & technology 41 2 632 - 8 2007年01月15日 [査読有り][通常論文]
     
    For more efficient control and prediction of membrane biofouling in membrane bioreactors (MBRs), a fundamental understanding of mechanisms of membrane biofouling is essential. In this study, we operated full-scale submerged MBRs using real municipal wastewater delivered from the primary sedimentation basin of a municipal wastewater treatment facility over 3 months, and the adhesion and formation of biofilms on 0.4-microm pore size polyethylene hollow-fiber microfiltration (MF) membrane surfaces, separated from simple deposition of sludge cake, were monitored using scanning electron microscopy (SEM). In addition, the compositions of planktonic and biofilm microbial communities in the MBR were analyzed using culture independent molecular-based methods (i.e., fluorescent in situ hybridization (FISH) and 16S rRNA gene sequence analysis). The SEM and LIVE/DEAD staining analyses clearly showed that the biofilms gradually developed on the membrane surfaces with time, which had a strong positive correlation with the increase in trans-membrane pressure (TMP). This indicated that the biofilm formation induced the membrane fouling. The FISH results revealed that the microbial communities on membrane surfaces were quite different from those in the planktonic biomass in the mixed liquor. Moreover, FISH and 16S rRNA gene sequence analyses revealed that a specific phylogenetic group of bacteria, the Betaproteobacteria, probably played a major role in development of the mature biofilms, which led to the severe irreversible membrane biofouling.
  • Ikuo Tsushima, Yuji Ogasawara, Masaki Shimokawa, Tomonori Kindaichi, Satoshi Okabe
    Water science and technology : a journal of the International Association on Water Pollution Research 55 8-9 9 - 17 2007年 [査読有り][通常論文]
     
    The anaerobic ammonium oxidation (Anammox) process is a new efficient and cost effective method of ammonium removal from wastewater. Under strictly anoxic condition, ammonium is directly oxidised with nitrite as electron acceptor to dinitrogen gas. However, it is extremely difficult to cultivate Anammox bacteria due to their low growth rate. This suggests that a rapid and efficient start-up of Anammox process is the key to practical applications. To screen appropriate seeding sludge with high Anammox potential, a real-time quantitative PCR assay with newly designed primers has been developed. Thereafter, the seeding sludge with high abundance of Anammox bacteria (1.7 x 10(8) copies/mg-dry weight) was selected and inoculated into an upflow anaerobic biofilters (UABs). The UABs were operated for more than 1 year and the highest nitrogen removal rate of 24.0 kg-N m-3 day(-1) was attained. In addition, the ecophysiology of Anammox bacteria (spatial distribution and in situ activity) in biofilms was analysed by combining a full-cycle 16S rRNA approach and microelectrodes. The microelectrode measurement clearly revealed that a successive vertical zonation of the partial nitrification (NH4+ to NO2-), Anammox reaction and denitrification was developed in the biofilm in the UAB. This result agreed with the spatial distribution of corresponding bacterial populations in the biofilm. We linked the micro-scale information (i.e. single cell and/or biofilm levels) with the macro-scale information (i.e. the reactor level) to understand the details of Anammox reaction occurring in the UABs.
  • Satoshi Okabe
    SOCIOLOGICAL THEORY AND METHODS 22 2 169 - 187 2007年 [査読有り][通常論文]
     
    Since 1990's in Japan, the increase of non-regular employment in youth has had a great impact on the way of getting their achievement of their social status. Ever since then, a lot of studies focusing on the differences among youth, such as income gap, have been provided, whereas studies about consciousness or sense of values of youth are relatively few. In this paper, I pay an attention to job satisfaction of Japanese youth and try to find determinant factors of their job satisfaction. As a result, job satisfaction of single male non-regular employees remains at the lowest level among young people. The youth belonging to this category has an affinity with a jobless youth on life awareness side, and would not have enough living or educational background. Focusing on one's past experience, I found that the experience to come in contact with adults is an essential factor of job satisfaction. To solve the issues of employment in youth, it is important not only to improve the work condition of youth, but also to promote the experience to come in contact with adults at a compulsory education stage.
  • Satoshi Okabe, Chieko Fuse, Shinji Sugihara, Sadahito Aoshima, Mitsuhiro Shibayama
    PHYSICA B-CONDENSED MATTER 385 756 - 758 2006年11月 [査読有り][通常論文]
     
    The structural transitions in block (Block) and gradient copolymer (Grad) aqueous solutions were investigated with small-angle neutron scattering and were compared to each other. The micelle formation was observed both in Grad and in Block systems. The micelle formation in Grad system was rather continuous, while that in Block underwent a stepwise transition. The size and its variation with temperature of the resulting micelles in Grad copolymer were also different from those in Block system. Possible transition mechanisms for these copolymer systems are described by taking account of the continuous variation in the composition along the polymer chain of Grad copolymer. (c) 2006 Elsevier B.V. All rights reserved.
  • PB-07 異種微生物間コミュニケーションによるPseudomonas aeruginosaの抗生物質耐性、病原性、遺伝子発現制御の発見(共生/相互作用,ポスターセッションB,(1)ポスター発表会,研究発表会)
    八幡, 穣, 岡部, 聡, 伊藤, 暁信, 間世田, 英明, 内山, 裕夫, 野村, 暢彦, NAKAJIMA, Toshiaki
    日本微生物生態学会講演要旨集 0 22 125  日本微生物生態学会 2006年10月 [査読無し][通常論文]
  • Tomonori Kindaichi, Yoshiko Kawano, Tsukasa Ito, Hisashi Satoh, Satoshi Okabe
    BIOTECHNOLOGY AND BIOENGINEERING 94 6 1111 - 1121 2006年08月 [査読有り][通常論文]
     
    Population dynamics of ammonia-oxidizing bacteria (AOB) and uncultured Nitrospira-like nitrite-oxidizing bacteria (NOB) dominated in autotrophic nitrifying biofilms were determined by using real-time quantitative polymerase chain reaction (RTQ-PCR) and fluorescence in situ hybridization (FISH). Although two quantitative techniques gave the comparable results the RTQ-PCR assay was easier and faster than the FISH technique for quantification of both nitrifying bacteria in dense microcolony-forming nitrifying biofilms. Using this RTQ-PCR assay, we could successfully determine the maximum specific growth rate (mu = 0.021/h) of uncultured Nitrospira-like NOB in the suspended enrichment culture. The population dynamics of nitrifying bacteria in the biofilm revealed that once they formed the biofilm, the both nitrifying bacteria grew slower than in planktonic cultures. We also calculated the spatial distributions of average specific growth rates of both nitrifying bacteria in the biofilm based on the concentration profiles of NH4+, NO2-, and O-2, which were determined by microelectrodes, and the double-Monod model. This simple model estimation could explain the stratified spatial distribution of AOB and Nitrospira-like NOB in the biofilm. The combination of culture-independent molecular techniques and microelectrode measurements is a very powerful approach to analyze the in situ kinetics and ecophysiology of nitrifying bacteria including uncultured Nitrospira-like NOB in complex biofilm communities. (c) 2006 Wiley Periodicals, Inc.
  • H Satoh, T Yamakawa, T Kindaichi, T Ito, S Okabe
    BIOTECHNOLOGY AND BIOENGINEERING 94 4 762 - 772 2006年07月 [査読有り][通常論文]
     
    The bacterial community structure, in situ spatial distributions and activities of nitrifying and denitrifying bacteria in biofilms treating industrial waste-water were investigated by combination of the 16S rRNA gene clone analysis, fluorescence in situ hybridization (FISH) and microelectrodes. These results were compared with the nitrogen removal capacity of the industrial wastewater treatment plant (IWTP). Both nitrification and denitrification occurred in the primary denitrification (PD) tank and denitrification occurred in the secondary denitrification (SD) tank. In contrast, nitrification and denitrification rates were very low in the nitrification (N) tank. 16S rRNA gene clone sequence analysis revealed that the bacteria affiliated with Alphaproteobacteria, followed by Betaproteobacteria, were numerically important microbial groups in three tanks. The many clones affiliated with Alphaproteobacteria were closely related to the denitrifying bacteria (e.g., Hyphomicrobium spp., Rhodopseudomonas palustris, and Rhodobacter spp.). In addition, Methylophilus leisingeri affiliated with Betaproteobacteria, which favorably utilized methanol, was detected only in the SD-tank to which methanol was added. Nitrosomonas europaea and Nitrosomonas marina were detected as the ammonia-oxidizing bacteria affiliated with Betaproteobacteria throughout this plant, although the dominant species of them was different among three tanks. Nitrifying bacteria were mainly detected in the upper parts of the PD-biofilm whereas their populations were low in the upper parts of the N-biofilm. The presence of denitrifying bacteria affiliated with Hyphomicrobium spp. in SD- and N-biofilms was verified by FISH analysis. Microelectrode measurements showed that the nitrifying bacteria present in the N- and PD-biofilms were active and the bacteria present in the SD-biofilm could denitrify. (c) 2006 Wiley Periodicals, Inc.
  • Olga Savichtcheva, Satoshi Okabe
    Water research 40 13 2463 - 76 2006年07月 [査読有り][通常論文]
     
    The ecological and survival characteristics of bacterial, viral and parasitic pathogens vary under environmental conditions, indicating that probably no single indicator organism can predict the presence of all enteric pathogens for all types of waters and different host-associated fecal pollution. If there are true correlations between indicator organisms and pathogens, it is necessary to find out to what extent and under which circumstances these organisms can be used as reliable indicators of fecal pollution. Application of conventional and alternative fecal indicators has greatly enhanced our abilities to predict and reduce health risk associated with the use of surface waters. New molecular-based techniques have shown that combined use of conventional and alternative indicators for fecal pollution increases both the detection sensitivity and specificity of fecal pollution and associated pathogens. In this review, we, therefore, summarize the advantages and limitations of conventional and alternative fecal indicators in terms of predicting pathogen presence as well as current and future methodologies for direct pathogen monitoring in environmental waters. This manuscript is mainly focused on the relationships between microbial fecal indicators and the presence of pathogens, which have not previously been summarized yet and could nicely supplement with recent literature reviews on microbial source tracking.
  • Yoshiyuki Nakamura, Hisashi Satoh, Tomonori Kindaichi, Satoshi Okabe
    Environmental science & technology 40 5 1532 - 9 2006年03月01日 [査読有り][通常論文]
     
    The community structure, spatial distributions, and in situ activity of ammonia-oxidizing bacteria (AOB) representing the Betaproteobacteria and nitrite-oxidizing bacteria (NOB) representing the genus Nitrospira in three different river sediments with different pollution sources and levels along the Niida River, Hachinohe, Japan, were investigated by the combined use of 16S rRNA gene-cloning analysis, real-time quantitative polymerase chain reaction (RTQ-PCR) assays, and microelectrodes. The goal of this research was to evaluate the contribution of nitrifying activity in the sediment to the overall nitrogen elimination rate in this river. The 16S rRNA gene-cloning analysis revealed that the community structures of AOB and Nitrospira-like NOB are present in three sediments. On the basis of the results of 16S rRNA gene-cloning analysis, the RTQ-PCR assay using a TaqMan probe was developed and optimized for the quantification of the Nitrospira-like NOB. In the sediments, AOB specific 16S rRNA genes were detected in the range of 10(6) to 10(7) copies/cm3 and evenly distributed over the sampled sediment depth (0-5 mm), whereas the Nitrospira-like NOB 16S rRNA gene copy numbers per cm3 were 1-2 orders of magnitude higher than the AOB copy numbers. Under light conditions, intensive oxygenic photosynthesis occurred in the surface and increased the maximal O2 concentration and O2 penetration depth in all sediments. This concomitantly stimulated nitrifying bacteria present in diurnally anoxic deeper zones and expanded nitrification zones, which consequently increased the total NH4+ consumption rate in the sediment (i.e., total NH4+ flux into the sediment). The results suggested that the in situ nitrifying activity was restricted mainly to the surface 2 mm of the sediment and linked with photosynthetic activity, which obviously plays an important role in nitrogen elimination in this river.
  • BE Rittmann, M Hausner, F Loffler, NG Love, G Muyzer, S Okabe, DB Oerther, J Peccia, L Raskin, M Wagner
    ENVIRONMENTAL SCIENCE & TECHNOLOGY 40 4 1096 - 1103 2006年02月 [査読有り][通常論文]
  • Olga Savichtcheva, Noriko Okayama, Tsukasa Ito, Satoshi Okabe
    Biotechnology and bioengineering 92 3 356 - 63 2005年11月05日 [査読有り][通常論文]
     
    To evaluate the viability and survival ability of fecal Bacteroides spp. in environmental waters, a fluorescence-based live/dead staining method using ViaGram Red+ Bacterial gram stain and viability kit was combined with fluorescent in situ hybridization (FISH) with 16S rRNA-targeted oligonucleotide probe (referred as LDS-FISH). The proposed LDS-FISH was a direct and reliable method to detect fecal Bacteroides cells and their viability at single-cell level in complex microbial communities. The pure culture of Bacteroides fragilis and whole human feces were dispersed in aerobic drinking water and incubated at different water temperatures (4 degrees C, 13 degrees C, 18 degrees C, and 24 degrees C), and then the viability of B. fragilis and fecal Bacteroides spp. were determined by applying the LDS-FISH. The results revealed that temperature and the presence of oxygen have significant effects on the survival ability. Increasing the temperature resulted in a rapid decrease in the viability of both pure cultured B. fragilis cells and fecal Bacteroides spp. The live pure cultured B. fragilis cells could be found at the level of detection in drinking water for 48 h of incubation at 24 degrees C, whereas live fecal Bacteroides spp. could be detected for only 4 h of incubation at 24 degrees C. The proposed LDS-FISH method should provide useful quantitative information on the presence and viability of Bacteroides spp., a potential alternative fecal indicator, in environmental waters.
  • PB-37 異種微生物間コミュニケーションによるPseudomonas aeruginosaの抗生物質耐性、病原性、遺伝子発現制御の発見(共生/相互作用,ポスターセッションB,ポスター発表)
    八幡, 穣, 岡部, 聡, 伊藤, 暁信, 間世田, 英明, 内山, 裕夫, 野村, 暢彦
    日本微生物生態学会講演要旨集 0 21 157  日本微生物生態学会 2005年10月 [査読無し][通常論文]
  • H Tanami, H Tsuda, S Okabe, T Iwai, K Sugihara, Imoto, I, J Inazawa
    LABORATORY INVESTIGATION 85 9 1118 - 1129 2005年09月 [査読有り][通常論文]
     
    The question of whether any genetic differences exist between primary and colorectal cancers (CRCs) and their metastatic foci is controversial. To look for genetic aberrations involved in metastasis of CRCs to the liver, we performed subtractive comparative genomic hybridization (CGH) experiments using paired samples from 20 CRC patients with primary tumors and synchronous or metachronous liver metastases. Relatively frequent gains in DNA copy number were detected at 6p, suggesting the presence of one or more metastasis-related genes in the region. Analysis of 11 CRC cell lines using array-based CGH (CGH-array) revealed one 6p candidate gene, CCND3. Quantitative reverse transcriptase-polymerase chain reaction experiments showed that CCND3 was significantly upregulated in liver-metastatic lesions compared with primary lesions (P<0.0152). In addition, immunohistochemical analysis of 120 primary CRC tumors demonstrated that cyclin D3 expression in the region of rolled edge was significantly associated with total recurrence, especially hematogenous recurrence ( P = 0.0307). The results implied involvement of cyclin D3 in liver metastasis of CRC, and the data may contribute to the development of a novel therapy or diagnostic agent for this currently intractable disease. Our experiments also confirmed the power of subtractive CGH and CGH-array analysis for identifying cancer-related genes.
  • H Satoh, Y Sasaki, Y Nakamura, S Okabe, T Suzuki
    BIOTECHNOLOGY AND BIOENGINEERING 91 2 133 - 138 2005年07月 [査読有り][通常論文]
     
    In order to assess the applicability of using microelectrodes as a tool for inhibition tests, temporal and spatial inhibitory effects of 2-chlorophenol (2-CP) on O-2 respiration and nitrification activities in municipal wastewater biofilms were investigated using microelectrodes for O-2 and NH4+. The time-course microelectrode measurements demonstrated that 2-CP inhibited O-2 respiration and nitrification activities within 6-18 min. The microbial activities were inhibited only in the upper 400 pm of the biofilms by 2-CP, and the bacteria present in the deeper parts of the biofilms were still active, probably due to limited penetration of 2-CP. These results could reasonably explain the difference in inhibitory ratios of the O-2 respiration and nitrification activities in the biofilms. O-2 respiration activity was incompletely inhibited, which was attributed to the presence Of O-2 respiration activities in the deeper parts of the biofilm. In contrast, nitrification activity was significantly inhibited because ammonia-oxidizing bacteria were present in the upper parts of the biofilm. These results indicate that the microelectrodes with a very quick response time and a high spatial resolution are useful tools to study temporal and spatial inhibitory effects of inhibitors on in situ microbial activities in biofilms. (c) 2005 Wiley Periodicals, Inc.
  • S Okabe, T Kindaichi, T Ito
    APPLIED AND ENVIRONMENTAL MICROBIOLOGY 71 7 3987 - 3994 2005年07月 [査読有り][通常論文]
     
    The cross-feeding of microbial products derived from C-14-labeled nitrifying bacteria to heterotrophic bacteria coexisting in an autotrophic nitrifying biofilm was quantitatively analyzed by using microautoradiography combined with fluorescence in situ hybridization (MAR-FISH). After only nitrifying bacteria were labeled with [C-14] bicarbonate, biofilm samples were incubated with and without NH4+ as a sole energy source for 10 days. The transfer of C-14 originally incorporated into nitrifying bacterial cells to heterotrophic bacteria was monitored with time by using MAR-FISH. The MAR-FISH analysis revealed that most phylogenetic groups of heterotrophic bacteria except the beta-Proteobacteria showed significant uptake of C-14-labeled microbial products. In particular, the members of the Chloroflexi were strongly MAR positive in the culture without NH4+ addition, in which nitrifying bacteria tended to decay. This indicated that the members of the Chloroflexi preferentially utilized microbial products derived from mainly biomass decay. On the other hand, the members of the Cytophaga-Flavobacterium cluster gradually utilized C-14-labeled products in the culture with NH4+ addition in which nitrifying bacteria grew. This result suggested that these bacteria preferentially utilized substrate utilization-associated products of nitrifying bacteria and/or secondary metabolites of C-14-labeled structural cell components. Our results clearly demonstrated that the coexisting heterotrophic bacteria efficiently degraded and utilized dead biomass and metabolites of nitrifying bacteria, which consequently prevented accumulation of organic waste products in the biofilm.
  • Satoshi Okabe, Tsukasa Ito, Kenichi Sugita, Hisashi Satoh
    Applied and environmental microbiology 71 5 2520 - 9 2005年05月 [査読有り][通常論文]
     
    The succession of sulfur-oxidizing bacterial (SOB) community structure and the complex internal sulfur cycle occurring in wastewater biofilms growing under microaerophilic conditions was analyzed by using a polyphasic approach that employed 16S rRNA gene-cloning analysis combined with fluorescence in situ hybridization, microelectrode measurements, and standard batch and reactor experiments. A complete sulfur cycle was established via S(0) accumulation within 80 days in the biofilms in replicate. This development was generally split into two phases, (i) a sulfur-accumulating phase and (ii) a sulfate-producing phase. In the first phase (until about 40 days), since the sulfide production rate (sulfate-reducing activity) exceeded the maximum sulfide-oxidizing capacity of SOB in the biofilms, H(2)S was only partially oxidized to S(0) by mainly Thiomicrospira denitirificans with NO(3)(-) as an electron acceptor, leading to significant accumulation of S(0) in the biofilms. In the second phase, the SOB populations developed further and diversified with time. In particular, S(0) accumulation promoted the growth of a novel strain, strain SO07, which predominantly carried out the oxidation of S(0) to SO(4)(2-) under oxic conditions, and Thiothrix sp. strain CT3. In situ hybridization analysis revealed that the dense populations of Thiothrix (ca. 10(9) cells cm(-3)) and strain SO07 (ca. 10(8) cells cm(-3)) were found at the sulfur-rich surface (100 microm), while the population of Thiomicrospira denitirificans was distributed throughout the biofilms with a density of ca. 10(7) to 10(8) cells cm(-3). Microelectrode measurements revealed that active sulfide-oxidizing zones overlapped the spatial distributions of different phylogenetic SOB groups in the biofilms. As a consequence, the sulfide-oxidizing capacities of the biofilms became high enough to completely oxidize all H(2)S produced by SRB to SO(4)(2-) in the second phase, indicating establishment of the complete sulfur cycle in the biofilms.
  • Tsukasa Ito, Kenichi Sugita, Isao Yumoto, Yoshinobu Nodasaka, Satoshi Okabe
    International journal of systematic and evolutionary microbiology 55 Pt 3 1059 - 1064 2005年05月 [査読有り][通常論文]
     
    A novel mesophilic, chemolithoautotrophic, sulfur-oxidizing bacterium, designated strain SO07(T), was isolated from a microaerobic waste-water biofilm. Chemolithoautotrophic growth was observed with elemental sulfur, sulfide and thiosulfate as sole electron donors and oxygen as electron acceptor. Anaerobic and heterotrophic growth were not observed. Nitrate was not used as a terminal electron acceptor. The optimum pH and temperature for growth were pH 7.5 and 30 degrees C, respectively. The major isoprenoid quinone was Q-8. The DNA G + C content of strain SO07(T) was 47.1 mol%. Phylogenetic analysis of 16S rRNA gene sequences demonstrated that strain SO07(T) formed a monophyletic group in the gamma-Proteobacteria with only 89 % similarity to members of the genus Halothiobacillus, its nearest phylogenetic neighbours. In addition, the isolate differed from members of the genus Halothiobacillus in its requirement for and tolerance of NaCl; strain SO07(T) was unable to grow in NaCl concentrations of more than 180 mM. On the basis of phylogenetic, chemotaxonomic and physiological data, it is proposed that isolate SO07(T) (=JCM 12417(T) = ATCC BAA-1033(T)) represents the type strain of a novel species in a new genus, Thiovirga sulfuroxydans gen. nov., sp. nov.
  • Tsukasa Ito, Kenichi Sugita, Satoshi Okabe
    Applied and environmental microbiology 70 5 3122 - 9 2004年05月 [査読有り][通常論文]
     
    We successfully isolated a novel aerobic chemolithotrophic sulfur-oxidizing bacterium, designated strain SO07, from wastewater biofilms growing under microaerophilic conditions. For isolation, the use of elemental sulfur (S(0)), which is the most abundant sulfur pool in the wastewater biofilms, as the electron donor was an effective measure to establish an enrichment culture of strain SO07 and further isolation. 16S rRNA gene sequence analysis revealed that newly isolated strain SO07 was affiliated with members of the genus Halothiobacillus, but it was only distantly related to previously isolated species (89% identity). Strain SO07 oxidized elemental sulfur, thiosulfate, and sulfide to sulfate under oxic conditions. Strain SO07 could not grow on nitrate. Organic carbons, including acetate, propionate, and formate, could not serve as carbon and energy sources. Unlike other aerobic sulfur-oxidizing bacteria, this bacterium was sensitive to NaCl; growth in medium containing more than 150 mM was negligible. In situ hybridization combined with confocal laser scanning microscopy revealed that a number of rod-shaped cells hybridized with a probe specific for strain SO07 were mainly present in the oxic biofilm strata (ca. 0 to 100 micro m) and that they often coexisted with sulfate-reducing bacteria in this zone. These results demonstrated that strain SO07 was one of the important sulfur-oxidizing populations involved in the sulfur cycle occurring in the wastewater biofilm and was primarily responsible for the oxidation of H(2)S and S(0) to SO(4)(2-) under oxic conditions.
  • Yoshiyuki Nakamura, Hisashi Satoh, Satoshi Okabe, Yoshimasa Watanabe
    Water research 38 9 2439 - 47 2004年05月 [査読有り][通常論文]
     
    The present study investigated photosynthetic rates and their regulation by light within the upper 5mm of sediment in a tidal area of Niida River in Hachinohe, Japan. Steady-state concentration profiles of O(2), NH(4)(+), NO(2)(-), H(2)S, and pH in the sediment were measured with microelectrodes. Microzonation of O(2) respiration, denitrification and SO(4)(2-) reduction was found in the sediment. When light intensities exceeded 1050 micromol photons/m(2)/s, net photosynthetic activity was detected in the upper 0.5mm of the microbial mat colonizing on the sediment surface in the tidal area. In contrast, gross photosynthetic activity was detected in the upper 1.0mm of the microbial mat at 1900 micromol photons/m(2)/s. As light intensity increased, the net photosynthetic rate and O(2) penetration depth increased. The maximal net photosynthetic rate and O(2) penetration depth were 6.1 micromol O(2)/cm(3)/h and 2.2mm, respectively, at 1900 micromol photons/m(2)/s. Net photosynthetic rates in the microbial mat in the tidal area were lower than in the upstream sediment. The analysis of continuous O(2) concentration measurements in different layers of the microbial mat during artificial light-dark cycles demonstrated that the photosynthetic activity response to changes in light intensity was extremely fast (a few seconds) and the O(2) concentration in the microbial mat became stable within 200s. The measurement of physical and chemical parameters in river water revealed that the study site was relatively polluted and sunlight intensity significantly fluctuated temporally. These results suggested that the in situ microbial processes occurring in the sediment fluctuated in accordance with periodic fluctuations in sunlight intensity.
  • Tomonori Kindaichi, Tsukasa Ito, Satoshi Okabe
    Applied and Environmental Microbiology 70 3 1641 - 1650 2004年03月 [査読有り][通常論文]
     
    Ecophysiological interactions between the community members (i.e., nitrifiers and heterotrophic bacteria) in a carbon-limited autotrophic nitrifying biofilm fed only NH4+ as an energy source were investigated by using a full-cycle 16S RRNA approach followed by microautoradiography (MAR)-fluorescence in situ hybridization (FISH). Phylogenetic differentiation (identification) of heterotrophic bacteria was performed by 16S rRNA gene sequence analysis, and FISH probes were designed to determine the community structure and the spatial organization (i.e., niche differentiation) in the biofilm. FISH analysis showed that this autotrophic nitrifying biofilm was composed of 50% nitrifying bacteria (ammonia-oxidizing bacteria [AOB] and nitrite-oxidizing bacteria [NOB]) and 50% heterotrophic bacteria, and the distribution was as follows: members of the alpha subclass of the class Proteobacteria (α-Proteobacteria), 23% γ-Proteobacteria, 13% green nonsulfur bacteria (GNSB), 9% Cytophaga-Flavobacterium-Bacteroides (CFB) division, 2% and unidentified (organisms that could not be hybridized with any probe except EUB338), 3%. These results indicated that a pair of nitrifiers (AOB and NOB) supported a heterotrophic bacterium via production of soluble microbial products (SMP). MAR-FISH revealed that the heterotrophic bacterial community was composed of bacteria that were phylogenetically and metabolically diverse and to some extent metabolically redundant, which ensured the stability of the ecosystem as a biofilm. α- and γ -Proteobacteria dominated the utilization of [14C]acetic acid and 14C-amino acids in this biofilm. Despite their low abundance (ca. 2%) in the biofilm community, members of the CFB cluster accounted for the largest fraction (ca. 64%) of the bacterial community consuming N-acetyl-D-[1- 14C]glucosamine (NAG). The GNSB accounted for 9% of the 14C-amino acid-consuming bacteria and 27% of the [ 14C]NAG-consuming bacteria but did not utilize [14C] acetic acid. Bacteria classified in the unidentified group accounted for 6% of the total heterotrophic bacteria and could utilize all organic substrates, including NAG. This showed that there was an efficient food web (carbon metabolism) in the autotrophic nitrifying biofilm community, which ensured maximum utilization of SMP produced by nitrifiers and prevented buildup of metabolites or waste materials of nitrifiers to significant levels.
  • H Satoh, H Ono, B Rulin, J Kamo, S Okabe, KI Fukushi
    WATER RESEARCH 38 6 1633 - 1641 2004年03月 [査読有り][通常論文]
     
    A membrane aerated biofilm reactor (MABR), in which O-2 was supplied from the bottom of the biofilm and NH4+ and organic carbon were supplied from the biofilm surface, was operated at different organic carbon loading rates and intra-membrane air pressures to investigate the occurrence of simultaneous chemical oxygen demand (COD) removal, nitrification and denitrification. The spatial distribution of nitrification and denitrification zones in the biofilms was measured with microelectrodes for O-2, NH4+, NO2-, NO3- and pH. When the MABR was operated at approximately 1.0 g-COD/m(2)/day of COD loading rate, simultaneous COD removal, nitrification and denitrification could be achieved. The COD loading rates and the intra-membrane air pressures applied in this study had no effect on the startup and the maximum rates of NH4+ oxidation in the MABRs. Microelectrode measurements showed that O-2 was supplied from the bottom of the MABR biofilm and penetrated the whole biofilm. Because the biofilm thickness increased during the operations, an anoxic layer developed in the upper parts of the mature biofilms while an oxic layer was restricted to the deeper parts of the biofilms. The development of the anoxic zones in the biofilms coincided with increase in the denitrification rates. Nitrification occurred in the zones from membrane surface to a point of ca. 60 mum. Denitrification mainly occurred just above the nitrification zones. The COD loading rates and the intra-membrane air pressures applied in this study had no effect on location of the nitrification and denitrification zones. (C) 2004 Elsevier Ltd. All rights reserved.
  • H Satoh, H Ono, B Rulin, J Kamo, S Okabe, KI Fukushi
    WATER RESEARCH 38 6 1633 - 1641 2004年03月 [査読無し][通常論文]
     
    A membrane aerated biofilm reactor (MABR), in which O-2 was supplied from the bottom of the biofilm and NH4+ and organic carbon were supplied from the biofilm surface, was operated at different organic carbon loading rates and intra-membrane air pressures to investigate the occurrence of simultaneous chemical oxygen demand (COD) removal, nitrification and denitrification. The spatial distribution of nitrification and denitrification zones in the biofilms was measured with microelectrodes for O-2, NH4+, NO2-, NO3- and pH. When the MABR was operated at approximately 1.0 g-COD/m(2)/day of COD loading rate, simultaneous COD removal, nitrification and denitrification could be achieved. The COD loading rates and the intra-membrane air pressures applied in this study had no effect on the startup and the maximum rates of NH4+ oxidation in the MABRs. Microelectrode measurements showed that O-2 was supplied from the bottom of the MABR biofilm and penetrated the whole biofilm. Because the biofilm thickness increased during the operations, an anoxic layer developed in the upper parts of the mature biofilms while an oxic layer was restricted to the deeper parts of the biofilms. The development of the anoxic zones in the biofilms coincided with increase in the denitrification rates. Nitrification occurred in the zones from membrane surface to a point of ca. 60 mum. Denitrification mainly occurred just above the nitrification zones. The COD loading rates and the intra-membrane air pressures applied in this study had no effect on location of the nitrification and denitrification zones. (C) 2004 Elsevier Ltd. All rights reserved.
  • Tomonori Kindaichi, Tsukasa Ito, Satoshi Okabe
    Applied and environmental microbiology 70 3 1641 - 50 2004年03月 [査読有り][通常論文]
     
    Ecophysiological interactions between the community members (i.e., nitrifiers and heterotrophic bacteria) in a carbon-limited autotrophic nitrifying biofilm fed only NH(4)(+) as an energy source were investigated by using a full-cycle 16S rRNA approach followed by microautoradiography (MAR)-fluorescence in situ hybridization (FISH). Phylogenetic differentiation (identification) of heterotrophic bacteria was performed by 16S rRNA gene sequence analysis, and FISH probes were designed to determine the community structure and the spatial organization (i.e., niche differentiation) in the biofilm. FISH analysis showed that this autotrophic nitrifying biofilm was composed of 50% nitrifying bacteria (ammonia-oxidizing bacteria [AOB] and nitrite-oxidizing bacteria [NOB]) and 50% heterotrophic bacteria, and the distribution was as follows: members of the alpha subclass of the class Proteobacteria (alpha-Proteobacteria), 23%; gamma-Proteobacteria, 13%; green nonsulfur bacteria (GNSB), 9%; Cytophaga-Flavobacterium-Bacteroides (CFB) division, 2%; and unidentified (organisms that could not be hybridized with any probe except EUB338), 3%. These results indicated that a pair of nitrifiers (AOB and NOB) supported a heterotrophic bacterium via production of soluble microbial products (SMP). MAR-FISH revealed that the heterotrophic bacterial community was composed of bacteria that were phylogenetically and metabolically diverse and to some extent metabolically redundant, which ensured the stability of the ecosystem as a biofilm. alpha- and gamma-Proteobacteria dominated the utilization of [(14)C]acetic acid and (14)C-amino acids in this biofilm. Despite their low abundance (ca. 2%) in the biofilm community, members of the CFB cluster accounted for the largest fraction (ca. 64%) of the bacterial community consuming N-acetyl-D-[1-(14)C]glucosamine (NAG). The GNSB accounted for 9% of the (14)C-amino acid-consuming bacteria and 27% of the [(14)C]NAG-consuming bacteria but did not utilize [(14)C]acetic acid. Bacteria classified in the unidentified group accounted for 6% of the total heterotrophic bacteria and could utilize all organic substrates, including NAG. This showed that there was an efficient food web (carbon metabolism) in the autotrophic nitrifying biofilm community, which ensured maximum utilization of SMP produced by nitrifiers and prevented buildup of metabolites or waste materials of nitrifiers to significant levels.
  • Satoshi Okabe, Tomonori Kindaichi, Tsukasa Ito, Hisashi Satoh
    Biotechnology and bioengineering 85 1 86 - 95 2004年01月05日 [査読有り][通常論文]
     
    A fine-scale in situ spatial organization of ammonia-oxidizing bacteria (AOB) in biofilms was investigated by combining molecular techniques (i.e., fluorescence in situ hybridization (FISH) and 16S rDNA-cloning analysis) and microelectrode measurements. Important parameters of AOB microcolonies such as size distribution and areal cell density of the microcolonies were determined and correlated with substrate microprofiles in the biofilms. In situ hybridization with a nested 16S rRNA-targeted oligonucleotide probe set revealed two different populations of AOB, Nitrosomonas europaea-lineage and Nitrosospira multiformis-lineage, coexisting in an autotrophic nitrifying biofilm. Nitrosospira formed looser microcolonies, with an areal cell density of 0.51 cells microm(-2), which was half of the cell density of Nitrosomonas (1.12 cells microm(-2)). It is speculated that the formation of looser microcolonies facilitates substrate diffusion into the microcolonies, which might be a survival strategy to low O(2) and NH(4) (+) conditions in the biofilm. A long-term experiment (4-week cultivation at different substrate C/N ratios) revealed that the size distribution of AOB microcolonies was strongly affected by better substrate supply due to shorter distance from the surface and the presence of organic carbon. The microcolony size was relatively constant throughout the autotrophic nitrifying biofilm, while the size increased by approximately 80% toward the depth of the biofilm cultured at the substrate C/N = 1. A short-term ( approximately 3 h) organic carbon addition experiment showed that the addition of organic carbon created interspecies competition for O(2) between AOB and heterotrophic bacteria, which dramatically decreased the in situ NH(4) (+)-uptake activity of AOB in the surface of the biofilms. This result might explain the spatial distribution of AOB microcolony size in the biofilms cultured at the substrate C/N = 1. These experimental results suggest O(2) and organic carbon were the main factors controlling the spatial organization and activity of AOB in biofilms. These findings are significantly important to further improve mathematical models used to describe how the slow-growing AOB develop their niches in biofilms and how that configuration affects nitrification performance in the biofilm.
  • Satoshi Okabe, Tomonori Kindaichi, Tsukasa Ito
    Microbes and Environments 19 2 83 - 98 2004年 [査読無し][通常論文]
     
    A major goal of microbial ecology is to study the abundance, localization, and activities of microorganisms in situ in order to understand ecophysiological roles that the microorganisms play in complex natural ecosystems. In fact, in typical microbial habitats such as biofilms, sediments, and microbial aggregates, resources and physicochemical conditions are dynamically changing with time and across even a very tiny distance because of metabolic activities and substrate transport limitation. To directly correlate microbial identity (16S rRNA-based phylogeny) to the specific metabolic function of individual cells within such complex and heterogeneous microbial habitats, several new molecular-based techniques have been developed in the last decade. These techniques exploit in situ simultaneous phylogenetic identification and metabolic capabilities of even uncultured microorganisms without the need to isolate them in culture. Microautoradiography is a powerful but “rather old” tool, with which the in situ uptake of specific radiolabeled substrates by individual cells can be determined. Fluorescence in situ hybridization (FISH) is a new molecular-based technique that allows the in situ phylogenetic identification of individual cells. However, FISH cannot provide sufficient information on metabolic capabilities, because phylogeny and phenotype are rarely congruent. Recently, microautoradiography and FISH have been successfully combined to further improve the complementary strengths of the two methods. Microautoradiography combined with FISH (MAR-FISH) can be used to simultaneously examine the phylogenetic identity and the relative or actual specific activity of microorganisms within a complex microbial community at a single-cell level. This article overviews the principle, experimental protocol and application of the MAR-FISH technique, as well as current developments of other analytical techniques for in situ microbial functions (metabolic activities) from a single-cell level to community levels. © 2004, Japanese Society of Microbial Ecology & The Japanese Society of Soil Microbiology. All rights reserved.
  • Okabe S, Satoh H, Ito T, Watanabe Y
    Journal of Water and Environment Technology 2 2 65 - 74 2004年 [査読無し][通常論文]
  • T Sato, K Yoshinaga, S Okabe, T Okawa, T Higuchi, M Enomoto, T Takizawa, K Sugihara
    JAPANESE JOURNAL OF CLINICAL ONCOLOGY 33 12 631 - 635 2003年12月 [査読有り][通常論文]
     
    Background: Cyclooxygenase (COX)-2 may be linked to carcinogenesis. In the previous study, we examined COX-2 expression immunohistochemically in 95 adenomas and reported a significant correlation between its expression and the grade of dysplasia. To clarify the correlation between COX-2 expression and cell proliferation, we investigated Ki-67 labeling index using immunohistochemistry and its correlation with COX-2 expression. Methods: Immunohistological staining for Ki-67 antigen was performed on 95 colorectal adenomas previously reported. Results: The Ki-67 labeling index was significantly higher in the high-COX-2 group than in the low-COX-2 and negative groups in adenomas with moderate (44.5 +/- 6.4% vs 33.0 +/- 2.6%, 39.0 +/- 6.2%; P = 0.01, P < 0.001, respectively) or severe dysplasia (47.2 +/- 7.6% vs 40.3 +/- 7.2%, 35.0 +/- 5.4%; P = 0.02, P = 0.005, respectively). There was no correlation between Ki-67 labeling index and COX-2 expression in mild dysplasia. Conclusions: These results suggest that COX-2 may play a causal role in cell proliferation in carcinogenesis.
  • H Satoh, Y Nakamura, H Ono, S Okabe
    BIOTECHNOLOGY AND BIOENGINEERING 83 5 604 - 607 2003年09月 [査読有り][通常論文]
     
    Simultaneous nitrification and denitrification (SND) was investigated in the single aeration tank of a municipal wastewater treatment plant. Microelectrode measurements and batch experiments were performed to test for the presence of SND. Microelectrodes recorded the presence of O-2 concentration gradients in individual activated sludge flocs. When the O-2 concentration in the bulk liquid was <45 μM, anoxic zones were detected within flocs with a larger diameter (approximately 3000 pm). The O-2 penetration depth in the floc was found to be dependent on the O-2 concentration in the bulk liquid. Nitrification was restricted to the oxic zones, whereas denitrification occurred mainly in the anoxic zones. The nitrification rate of the activated sludge increased with increasing O-2 concentration in the bulk liquid, up to 40 μM, and remained constant thereafter. SND was observed in the aerated activated sludge when O-2 concentration was in the range of 10 to 35 μM. (C) 2003 Wiley Periodicals, Inc.
  • GH Chen, MT Wong, S Okabe, Y Watanabe
    WATER RESEARCH 37 13 3125 - 3135 2003年07月 [査読有り][通常論文]
     
    Dynamic response of nitrifying activated sludge batch cultures to increased chloride concentration was studied in this paper, which focused upon the changes in the specific nitrification rate (SNR) and nitrifier population when the chloride level was gradually or stepwise increased to 30,000 mg Cl L-1. The dominant species of ammonia-oxidizers and nitrite-oxidizers in the population were examined by Fluorescent in situ hybridization technique with 16S rRNA-targeted oligonucleotide probes. It was found that neither chloride increasing approaches affected the SNR of the batch cultures before the chloride concentration exceeded 10,000 mg Cl L-1, after which the stepwise increase approach reduced the SNR more significantly than the gradual increase approach. From 10,000 to 18,000 mg Cl L-1 a down-and-up pattern of the SNR variation appeared in both approaches, which was associated with the change in the dominant species of ammonia-oxidizers from non-saline-resistant species such as Nitrosomonas europaea-lineage and Nitrosomonas eutropha to saline-resistant species, such as the Nitrosococcus mobilis-lineage. Nitrobacter was the only dominant species when the chloride concentration was below 10,000 mg Cl L-1, where no nitrite-oxidizers survived. Therefore, the 10,000 mg Cl L-1 chloride level is a critical level for the shift of the nitrifier population in the nitrifying activated sludge batch cultures. (C) 2003 Elsevier Science Ltd. All rights reserved.
  • H Satoh, S Okabe, Y Yamaguchi, Y Watanabe
    WATER RESEARCH 37 9 2206 - 2216 2003年05月 [査読有り][通常論文]
     
    Three rotating disk biofilm reactors were operated to evaluate whether bioaugmentation and biostimulation can be used to improve the start-up of microbial nitrification. The first reactor was bioaugmented during start-up period with an enrichment culture of nitrifying bacteria, the second reactor received a synthetic medium containing NH4+ and NO2- to facilitate concomitant proliferation of ammonia- and nitrite-oxidizing bacteria, and the third reactor was used as a control. To evaluate the effectiveness of bioaugmentation and biostimulation approaches, time-dependent developments of nitrifying bacterial community and in situ nitrifying activity in biofilms were monitored by fluorescence in situ hybridization (FISH) technique and microelectrode measurements of NH4+, NO2-, NO3-, and O-2. In situ hybridization results revealed that addition of the enrichment culture of nitrifying bacteria significantly facilitated development of dense nitrifying bacterial populations in the biofilm shortly after, which led to a rapid start-up and enhancement of in situ nitrification activity. The inoculated bacteria could proliferate and/or survive in the biofilm. In addition, the addition of nitrifying bacteria increased the abundance of nitrifying bacteria in the surface of the biofilm, resulting in the higher nitrification rate. On the other hand, the addition of 2.1 mM NO2- did not stimulate the growth of nitrite-oxidizing bacteria and did inhibit the proliferation of ammonia-oxidizing bacteria instead. Thus, the start-up of NO2- oxidation was unchanged, and the start-up of NH4+ oxidation was delayed. In all the three biofilm reactors, data sets of time series analyses on population dynamics of nitrifying bacteria determined by FISH, in situ nitrifying activities determined by microelectrode measurements, and the reactor performances revealed an approximate agreement between the appearance of nitrifying bacteria and the initiation of nitrification activity, suggesting that the combination of these techniques was a very powerful monitoring tool to evaluate the effectiveness of bioaugmentation and biostimulation strategies. (C) 2002 Elsevier Science Ltd. All rights reserved.
  • Satoshi Okabe, Cecilia M Santegoeds, Dirk De Beer
    Biotechnology and bioengineering 81 5 570 - 7 2003年03月05日 [査読有り][通常論文]
     
    Microelectrode, fluorescence in situ hybridization (FISH), and denaturing gradient gel electrophoresis (DGGE) analyses were used to investigate the effect of nitrite and nitrate on in situ sulfide production in an activated sludge immobilized agar gel film. Microelectrode measurements of O(2), H(2)S, NO(3)(-), NO(2)(-), and pH revealed that the addition of NO(2)(-) and NO(3)(-) forced sulfate reduction zones deeper in the agar gel and significantly reduced the in situ sulfide production levels. The sulfate reduction zone was consequently separated from O(2) and NO(2)(-) or NO(3)(-) respiration zones with increasing the concentrations of NO(2)(-) and NO(3)(-). These NO(2)(-) and NO(3)(-) treatments had only a transient effect on sulfide production. The in situ sulfide production quickly recovered to the previous levels when NO(2)(-) and NO(3)(-) were removed. The PCR-DGGE and FISH analyses revealed that 2-day-continuous addition of 500 microM NO(3)(-) did not change the metabolically active sulfate-reducing bacterial (SRB) community. On the basis of these data, it could be concluded that the addition of NO(2)(-) and NO(3)(-) did not kill SRB, but induced the interspecies competition for common carbon source (i.e., acetate) between nitrate-reducing heterotrophic bacteria and SRB and enhanced the oxidation of the produced sulfide, which were main possible causes of the suppression of in situ sulfide production in the agar gel.
  • Aurelio M Briones, Satoshi Okabe, Yoshiaki Umemiya, Niels-Birger Ramsing, Wolfgang Reichardt, Hidetoshi Okuyama
    Applied and environmental microbiology 68 6 3067 - 75 2002年06月 [査読有り][通常論文]
     
    Comparisons of the activities and diversities of ammonia-oxidizing bacteria (AOB) in the root environment of different cultivars of rice (Oryza sativa L.) indicated marked differences despite identical environmental conditions during growth. Gross nitrification rates obtained by the 15N dilution technique were significantly higher in a modern variety, IR63087-1-17, than in two traditional varieties. Phylogenetic analysis based on the ammonium monooxygenase gene (amoA) identified strains related to Nitrosospira multiformis and Nitrosomonas europaea as the predominant AOB in our experimental rice system. A method was developed to determine the abundance of AOB on root biofilm samples using fluorescently tagged oligonucleotide probes targeting 16S rRNA. The levels of abundance detected suggested an enrichment of AOB on rice roots. We identified 40 to 69% of AOB on roots of IR63087-1-17 as Nitrosomonas spp., while this subpopulation constituted 7 to 23% of AOB on roots of the other cultivars. These results were generally supported by denaturing gradient gel electrophoresis of the amoA gene and analysis of libraries of cloned amoA. In hydroponic culture, oxygen concentration profiles around secondary roots differed significantly among the tested rice varieties, of which IR63087-1-17 showed maximum leakage of oxygen. The results suggest that varietal differences in the composition and activity of root-associated AOB populations may result from microscale differences in O2 availability.
  • Satoshi Okabe, Cecilia M Santegoeds, Yoshimasa Watanabe, Dirk de Beer
    Biotechnology and bioengineering 78 2 119 - 30 2002年04月20日 [査読有り][通常論文]
     
    A combination of fluorescence in situ hybridization (FISH), microprofiles, and denaturing gradient gel electrophoresis (DGGE) analysis of PCR-amplified 16S rDNA fragments followed by hybridization analysis with specific probes was applied to investigate successional development of sulfate-reducing bacteria (SRB) community structure and in situ sulfide production activity within an activated sludge immobilized agar gel film. In this model biofilm system, since biases arising from biofilm heterogeneity can be ignored, the population dynamics of SRB in the agar gel is directly related to physiological capability and in situ activity of SRB. Microelectrode measurements showed that an anoxic zone was already developed at the beginning (0 day), a first sulfide production of 0.054 mumol H2S m(-2) x s(-1) was detected during the first week, and the rate increased gradually to 0.221 mumol H2S m(-2) x s(-1) in the fifth week. The most active sulfide production zone moved upward to the chemocline and intensified with time to form a narrow zone with high volumetric sulfide production rates. This result coincided with the shift of the spatial distributions of SRB populations determined by FISH. In situ hybridization with probe SRB385 for mainly general SRB of the delta Proteobacteria plus some gram-positive bacteria and probe 660 for Desulfobulbus indicated that the most abundant populations of SRB were primarily restricted to near the oxic/anoxic interface (chemocline). A close observation of the development of the vertical distributions of SRB populations revealed that the cell numbers of Desulfobulbus tripled (from 0.5 x 10(8) to 1.5 x 10(8) cells cm(-3)) near the oxic/anoxic interface. Similar growth (from 1.0 x10(8) to 4.5 x 10(8) cells cm(-3)) of Desulfovibrio-like SRB that hybridized with probe SRB385 was observed. PCR-DGGE followed by hybridization analysis revealed that one Desulfobulbus strain was detected from the beginning, and another strain appeared after 1 week, coinciding with the first detected sulfide production. In addition, three strains hybridizing with probe 687 (possibly Desulfovibrio) were also dominant SRB in the agar gel.
  • Tsukasa Ito, Satoshi Okabe, Hisashi Satoh, Yoshimasa Watanabe
    Applied and environmental microbiology 68 3 1392 - 402 2002年03月 [査読有り][通常論文]
     
    A combination of fluorescence in situ hybridization, microprofiles, denaturing gradient gel electrophoresis of PCR-amplified 16S ribosomal DNA fragments, and 16S rRNA gene cloning analysis was applied to investigate successional development of sulfate-reducing bacteria (SRB) community structure and in situ sulfide production activity within a biofilm growing under microaerophilic conditions (dissolved oxygen concentration in the bulk liquid was in the range of 0 to 100 microM) and in the presence of nitrate. Microelectrode measurements showed that oxygen penetrated 200 microm from the surface during all stages of biofilm development. The first sulfide production of 0.32 micromol of H(2)S m(-2) s(-1) was detected below ca. 500 microm in the 3rd week and then gradually increased to 0.70 micromol H(2)S m(-2) s(-1) in the 8th week. The most active sulfide production zone moved upward to the oxic-anoxic interface and intensified with time. This result coincided with an increase in SRB populations in the surface layer of the biofilm. The numbers of the probe SRB385- and 660-hybridized SRB populations significantly increased to 7.9 x 10(9) cells cm(-3) and 3.6 x 10(9) cells cm(-3), respectively, in the surface 400 microm during an 8-week cultivation, while those populations were relatively unchanged in the deeper part of the biofilm, probably due to substrate transport limitation. Based on 16S rRNA gene cloning analysis data, clone sequences that related to Desulfomicrobium hypogeium (99% sequence similarity) and Desulfobulbus elongatus (95% sequence similarity) were most frequently found. Different molecular analyses confirmed that Desulfobulbus, Desulfovibrio, and Desulfomicrobium were found to be the numerically important members of SRB in this wastewater biofilm.
  • Tsukasa Ito, Jeppe L Nielsen, Satoshi Okabe, Yoshimasa Watanabe, Per H Nielsen
    Applied and environmental microbiology 68 1 356 - 64 2002年01月 [査読有り][通常論文]
     
    We simultaneously determined the phylogenetic identification and substrate uptake patterns of sulfate-reducing bacteria (SRB) inhabiting a sewer biofilm with oxygen, nitrate, or sulfate as an electron acceptor by combining microautoradiography and fluorescent in situ hybridization (MAR-FISH) with family- and genus-specific 16S rRNA probes. The MAR-FISH analysis revealed that Desulfobulbus hybridized with probe 660 was a dominant SRB subgroup in this sewer biofilm, accounting for 23% of the total SRB. Approximately 9 and 27% of Desulfobulbus cells detected with probe 660 could take up [(14)C]propionate with oxygen and nitrate, respectively, as an electron acceptor, which might explain the high abundance of this species in various oxic environments. Furthermore, more than 40% of Desulfobulbus cells incorporated acetate under anoxic conditions. SRB were also numerically important members of H(2)-utilizing and (14)CO(2)-fixing microbial populations in this sewer biofilm, accounting for roughly 42% of total H(2)-utilizing bacteria hybridized with probe EUB338. A comparative 16S ribosomal DNA analysis revealed that two SRB populations, related to the Desulfomicrobium hypogeium and the Desulfovibrio desulfuricans MB lineages, were found to be important H(2) utilizers in this biofilm. The substrate uptake characteristics of different phylogenetic SRB subgroups were compared with the characteristics described to date. These results provide further insight into the correlation between the 16S rRNA phylogenetic diversity and the physiological diversity of SRB populations inhabiting sewer biofilms.
  • S OKABE, WL JONES, W LEE, WG CHARACKLIS
    BIOFOULING AND BIOCORROSION IN INDUSTRIAL WATER SYSTEMS 189 - 204 1994年 [査読有り][通常論文]

MISC

  • N'Dah Joel Koffi, Satoshi Okabe Chemical Engineering Journal 380 2020年01月15日 [査読無し][通常論文]
     
    © 2019 Elsevier B.V. Microbial fuel cells (MFCs) exhibit high capital cost and low energy output, which are major issues of the expected practical application of MFCs in wastewater treatment. Here, we have developed serpentine up-flow MFCs equipped with polyvinylidene fluoride (PVDF)-based activated carbon (AC) air-cathode (MFC-PVDF/AC) and continuously operated for more than 6 months with real domestic wastewater as a substrate. The MFC-PVDF/ACs achieved average total COD removal rates (5.11 ± 0.94 kg tCOD/m3/d) and power densities (3.96 ± 3.01 W/m3) without major water leakage, which were even higher than those of MFCs equipped with Pt-based air-cathode (MFC-Pts). The MFC-PVDF/ACs also achieved high and stable suspended solid (SS) removal efficiency (>90%) at 1.5-h HRT without any clogging event during the entire operation period. Since the PVDF-based AC air-cathode is less expensive, more durable, and easy to manufacture, expensive Pt cathodes are not necessary for MFCs treating low strength domestic wastewater. Furthermore, we have developed a low voltage booster (LVB) to increase low output voltage of MFC-PVDF/ACs (i.e., <0.4 V). Connecting a single LVB with a single MFC-PVDF/AC increased the voltage from <0.4 V to 4.35–5.2 V without the voltage reversal, which was enough to turn on three LED bulbs for >12 days. Taken together, the MFC-PVDF/AC with a LVB circuit exhibits excellent and stable performance of domestic wastewater treatment and usable power generation, suggesting that it could be used as a cost- and energy-saving primary wastewater treatment system.
  • Masaaki Kitajima, Satoshi Ishii, Tatsuma Takagi, Satoshi Okabe Microbiology resource announcements 8 (23) 2019年06月06日 [査読有り][通常論文]
     
    The Shigella bacterium is one of the most significant causes of waterborne and foodborne bacterial dysentery. A lytic bacteriophage infecting Shigella flexneri was isolated from wastewater in Japan. We report here the complete genome sequence of this bacteriophage, revealing that it belongs to the Myoviridae family and possesses linear genomic DNA.
  • Tsuyoshi Kato, Ayano Kobayashi, Wakana Oishi, Syun-Suke Kadoya, Satoshi Okabe, Naoya Ohta, Mohan Amarasiri, Daisuke Sano Journal of water and health 17 (3) 404 -415 2019年06月 [査読有り][通常論文]
     
    This study presents a novel methodology for estimating the concentration of environmental pollutants in water, such as pathogens, based on environmental parameters. The scientific uniqueness of this study is the prevention of excess conformity in the model fitting by applying domain knowledge, which is the accumulated scientific knowledge regarding the correlations between response and explanatory variables. Sign constraints were used to express domain knowledge, and the effect of the sign constraints on the prediction performance using censored datasets was investigated. As a result, we confirmed that sign constraints made prediction more accurate compared to conventional sign-free approaches. The most remarkable technical contribution of this study is the finding that the sign constraints can be incorporated in the estimation of the correlation coefficient in Tobit analysis. We developed effective and numerically stable algorithms for fitting a model to datasets under the sign constraints. This novel algorithm is applicable to a wide variety of the prediction of pollutant contamination level, including the pathogen concentrations in water.
  • Koji Matsunaga, Yu Okuyama, Reiko Hirano, Satoshi Okabe, Masahiro Takahashi, Hisashi Satoh Chemosphere 224 538 -543 2019年06月 [査読有り][通常論文]
     
    A simple analytical method was developed to determine the arsenite (As(III)) concentration using a DNA aptamer and gold nanoparticles (AuNPs). Prior to sample measurements, the method sensing mechanism was confirmed by analyzing the particle size of the AuNPs at each step of the analysis procedure, and the key operational parameters that affect the method performance were optimized. The optimal final NaCl concentration, incubation time with NaCl and pH of a 3-(N-morpholino) propanesulfonic acid buffer were 60 mM, 10 min and 7.3, respectively. A calibration curve was created under optimized operational conditions. The calibration curve was linear from a 1.0- to 10-μM As(III) concentration. The detection limit was 2.1 μM (161 μg/L). Using the calibration curve, we evaluated groundwater samples spiked with As(III). As(III) concentrations in groundwater pretreated with a 0.2-μm-pore-size membrane filter and cation-exchange resin were determined by using the method, which suggests that the proposed method can be used to determine the As(III) concentration in groundwater.
  • Mohan Amarasiri, Hiroki Kawai, Masaaki Kitajima, Satoshi Okabe, Daisuke Sano Water science and technology : a journal of the International Association on Water Pollution Research 79 (2) 342 -348 2019年01月 [査読有り][通常論文]
     
    Contribution of specific interactions between human enteric viruses and wastewater suspended solids on human enteric virus removal by microfiltration was studied. A cross-flow microfiltration system was used with rotavirus HAL1166 and Enterobacter cloacae SENG-6 as the model virus and wastewater suspended solid. Cleavage of rotavirus HAL1166 protein VP4 by trypsin produces the VP8* subunit, which specifically interacts with histo-blood group antigen (HBGA). In the presence of Enterobacter cloacae SENG-6, the trypsin-treated rotavirus concentration reduced with time (R2 > 0.6) compared to the reduction of non-trypsin treated rotavirus. Calculation of the gel/cake layer deposited on the membrane, consisting of Enterobacter cloacae SENG-6 and either trypsin-treated or non-trypsin treated rotavirus HAL1166, revealed that the microflocs consisting of trypsin-treated rotavirus and Enterobacter cloacae SENG-6 have lower porosity and permeability, displaying higher resistance to virus passage through the membrane. The results provide evidence that specific wastewater suspended solids-human enteric virus interaction can contribute to increasing the removal of human enteric viruses by microfiltration.
  • Godwin E. Oyiwona, James C. Ogbonna, Chukwudi Uzoma Anyanwu, Satoshi Okabe Bioresources and Bioprocessing 5 2018年12月01日 [査読無し][通常論文]
     
    © 2018, The Author(s). Background: Poultry droppings from poultry farms and rice husks obtained from rice milling process are generally considered as wastes and discarded in Nigeria. Although many studies have shown that microbial fuel cells (MFCs) can generate electricity from organic wastes, little or no study have examined MFCs for generating electricity from poultry droppings and rice husk as electrode material. Findings: Laboratory-scale double-chamber MFCs were inoculated with concentrations of poultry droppings wastewater and supplied with rice husk charcoal as anode and cathode electrodes for electricity generation. Power outputs and dissolved organic carbon (DOC) removal efficiencies were compared between MFCs using rice husk charcoal (RHCE) as electrode and those using carbon cloth (CCE) as electrodes. The RHCE-MFC 2 containing 477 mg L−1dissolved organic carbon produced a volumetric power density of 6.9 ± 3.1 W m−3which was higher than the control and the CCE-MFCs by a factor of 2 and achieved at DOC removal efficiencies of 40 ± 1.2%. Conclusions: The results suggest that poultry droppings wastewater is a feasible feedstock for generating electricity in MFCs. The findings also suggest that rice husk charcoal is a potentially useful electrode material in MFCs. [Figure not available: see fulltext.].
  • D. Sano, M. Tazawa, M. Inaba, S. Kadoya, R. Watanabe, T. Miura, M. Kitajima, S. Okabe Journal of Applied Microbiology 124 (4) 1001 -1007 2018年04月01日 [査読無し][通常論文]
     
    Aims: Cellular responses of an established cell line from human intestinal epithelial cells (INT-407 cells) against poliovirus (PV) infections were investigated in order to find cellular genetic markers for infectious PV detection. Methods and Results: Gene expression profile of INT-407 cells was analysed by DNA microarray technique when cells were infected with poliovirus 1 (PV1) (sabin) at multiplicity of infection of 10−3 and incubated for 12 h. Poliovirus infection significantly altered the gene expressions of two ion channels, KCNJ4 and SCN7A. The expression profile of KCNJ4 gene was further investigated by real-time RT-qPCR, and it was found that KCNJ4 gene was significantly regulated at 24 h postinfection of PV1. Conclusions: KCNJ4 gene, coding a potassium channel protein, is proposed as a cellular genetic marker for infectious PV detection. Significance and Impact of the Study: This is the first study to show the availability of cellular responses to detect infectious PV. The selection of cellular genetic markers for infectious viruses using DNA microarray and RT-qPCR can be applicable for the other enteric viruses.
  • 石丸美穂, 押木守, 荒木信夫, 岡部聡, 幡本将史, 山口隆司 日本水環境学会年会講演集 52nd 656 2018年03月09日 [査読無し][通常論文]
  • 小林香苗, 眞壁明子, 押木守, 金田一智規, 岡部聡 日本水環境学会年会講演集 52nd 379 2018年03月09日 [査読無し][通常論文]
  • So Ishizaki, Rimana Islam Papry, Hiroshi Miyake, Yuko Narita, Satoshi Okabe Frontiers in microbiology 9 3284 -3284 2018年 [査読有り][通常論文]
     
    Integrated microbial fuel cell (MFC) and membrane bioreactor (MBR) systems are a promising cost-effective and energy-saving technology for wastewater treatment. Membrane fouling is still an important issue of such integrated systems in which aeration (oxygen) is replaced with anode electrodes (anodic respiration). Here, we investigated the effect of culture conditions on the membrane fouling potential of fouling-causing bacteria (FCB). In the present study, Klebsiella quasipneumoniae strain S05, which is an exoelectrogenic FCB isolated from a MBR treating municipal wastewater, was cultured with different external electron acceptors (oxygen, nitrate, and solid-state anode electrode). As results, the fouling potential of S05 was lowest when cultured with anode electrode and highest without any external electron acceptor (p < 0.05, respectively). The composition of soluble microbial products (SMP) and extracellular polymeric substances (EPS) was also dependent on the type of electron acceptor. Protein and biopolymer contents in SMP were highly correlated with the fouling potential (R2 = 0.73 and 0.81, respectively). Both the fouling potential and yield of protein and biopolymer production were significantly mitigated by supplying electron acceptors sufficiently regardless of its types. Taken together, the aeration of MBR could be replaced with solid-state anode electrodes without enhancement of membrane fouling, and the anode electrodes must be placed sufficiently to prevent the dead spaces in the integrated reactor.
  • Seiya Hirano, Daisuke Sano, Satoshi Okabe, Masaaki Kitajima Noroviruses: Outbreaks, Control and Prevention Strategies 223 -242 2017年01月01日 [査読無し][通常論文]
     
    © 2017 Nova Science Publishers, Inc. Aptamer has been attracting much attention as a new biomolecular tool for viral diagnostic and therapeutic applications. Aptamer is a functional nucleic acid that folds into a unique three-dimensional conformation that binds to a specific target, which can be an alternative for antibodies. Rather, aptamer offers various advantages over antibodies as a target-specific recognition molecule: high affinity, specificity, and thermostability, low production cost, etc. Several studies have reported DNA aptamers for certain human norovirus strains as well as murine norovirus, which has enabled development and evaluation of aptasensor (aptamer-based biosensor) technology for rapid and sensitive norovirus detection. Therapeutic application of aptamer to inhibiting norovirus infections has not been clearly demonstrated to date, but there have been a few reports suggesting the potential of aptamer-based therapeutics. Taken together, it has been implied that aptamer is a new target-specific biomolecular tool that offers various potential applications to norovirus research, diagnosis, and therapeutics, which should contribute to the infection and disease control of norovirus by rapidly identifying potential infection routes and reducing the disease burden through therapeutic approach. Herein we summarize recent development and application of norovirus-specific aptamers and discuss future research directions towards better control of norovirus transmission/infections using this aptamer technology.
  • 押木守, 岡部聡 日本微生物生態学会大会(Web) 2017 ROMBUNNO.S‐070 (WEB ONLY) 2017年 [査読無し][通常論文]
  • Satoshi Ishii, Mohan Amarasiri, Satoshi Hashiba, Peiyi Yang, Satoshi Okabe, Daisuke Sano Genome Announcements 4 (4) e00893 -16 2016年08月 [査読無し][通常論文]
     
    © 2016 Ishii et al. Enterobacter sp. strain SENG-6, isolated from healthy human feces, produces histo-blood group antigen (HBGA)-like substances that can bind with human noroviruses. Based on the genome sequence analysis, strain SENG-6 belongs to the species Enterobacter cloacae. The genome sequence of this strain should help identify genes associated with the production of HBGA-like substances.
  • 小林香苗, 押木守, 金田一智規, 眞壁明子, 岡部聡 日本水環境学会年会講演集 50th 664 2016年03月10日 [査読無し][通常論文]
  • 中村新, 渡辺幸三, 八重樫咲子, 岡部聡, 中込とよ子, 中込治, 佐野大輔 日本水環境学会年会講演集 50th 398 2016年03月10日 [査読無し][通常論文]
  • Muhammad Ali, Mohamed Fauzi Haroon, Yuko Narita, Lei Zhang, Dario Rangel Shaw, Satoshi Okabe, Pascal E. Saikaly Genome Announcements 4 (6) 2016年 [査読無し][通常論文]
     
    The anaerobic ammonium-oxidizing (anam mox) bacterium "Candidatus Brocadia sp. 40" demonstrated the fastest growth rate compared to others in this taxon. Here, we report the 2.93-Mb draft genome sequence of this bacterium, which has 2,565 genecoding regions, 41 tRNAs, and a single rrn operon.
  • T. Ito, T. Kato, K. Takagishi, S. Okabe, D. Sano WATER SCIENCE AND TECHNOLOGY 72 (10) 1789 -1795 2015年11月 [査読無し][通常論文]
     
    Left-censored datasets of virus density in wastewater samples make it difficult to evaluate the virus removal efficiency in wastewater treatment processes. In the present study, we modeled the probabilistic distribution of virus removal efficiency in a wastewater treatment process with a Bayesian approach, and investigated how many detect samples in influent and effluent are necessary for accurate estimation. One hundred left-censored data of virus density in wastewater (influent and effluent) were artificially generated based on assumed log-normal distributions and the posterior predictive distribution of virus density, and the log-ratio distribution were estimated. The estimation accuracy of distributions was quantified by Bhattacharyya coefficient. When it is assumed that the accurate estimation of posterior predictive distributions is possible when a 100% positive rate is obtained for 12 pairs of influent and effluent, 11 out of 144, 60 out of 324, and 201 out of 576 combinations of detect samples gave an accurate estimation at the significant level of 0.01 in a Kruskal-Wallis test when the total sample number was 12, 18, and 24, respectively. The combinations with the minimum number of detect samples were (12, 9), (16, 10), and (21, 8) when the total sample number was 12, 18, and 24, respectively.
  • 押木守, 荒木信夫, 岡部聡 日本微生物生態学会大会(Web) 30th PD‐051 (WEB ONLY) 2015年 [査読無し][通常論文]
  • 富岡哲史, 渡辺幸三, 岡部聡, 佐野大輔 土木学会北海道支部論文報告集(CD-ROM) (70) ROMBUNNO.G-01 2014年02月 [査読無し][通常論文]
  • 押木守, 水戸佳祐, 木村善一郎, 金田一智規, 佐藤久, 岡部聡 日本ゲノム微生物学会年会要旨集 8th 69 2014年 [査読無し][通常論文]
  • R. M. L. D. Rathnayake, Y. Song, A. Tumendelger, M. Oshiki, S. Ishii, H. Satoh, S. Toyoda, N. Yoshida, S. Okabe WATER RESEARCH 47 (19) 7078 -7086 2013年12月 [査読無し][通常論文]
     
    Emission of nitrous oxide (N2O) during biological wastewater treatment is of growing concern since N2O is a major stratospheric ozone-depleting substance and an important greenhouse gas. The emission of N2O from a lab-scale granular sequencing batch reactor (SBR) for partial nitrification (PN) treating synthetic wastewater without organic carbon was therefore determined in this study, because PN process is known to produce more N2O than conventional nitrification processes. The average N2O emission rate from the SBR was 0.32 +/- 0.17 mgN L-1 h(-1), corresponding to the average emission of N2O of 0.8 +/- 0.4% of the incoming nitrogen load (1.5 +/- 0.8% of the converted NI-14"). Analysis of dynamic concentration profiles during one cycle of the SBR operation demonstrated that N2O concentration in off-gas was the highest just after starting aeration whereas N2O concentration in effluent was gradually increased in the initial 40 min of the aeration period and was decreased thereafter. Isotopomer analysis was conducted to identify the main N2O production pathway in the reactor during one cycle. The hydroxylamine (NH2OH) oxidation pathway accounted for 65% of the total N2O production in the initial phase during one cycle, whereas contribution of the NO reduction pathway to N2O production was comparable with that of the NH2OH oxidation pathway in the latter phase. In addition, spatial distributions of bacteria and their activities in single microbial granules taken from the reactor were determined with microsensors and by in situ hybridization. Partial nitrification occurred mainly in the oxic surface layer of the granules and ammonia-oxidizing bacteria were abundant in this layer. N2O production was also found mainly in the oxic surface layer. Based on these results, although N2O was produced mainly via NH2OH oxidation pathway in the autotrophic partial nitrification reactor, N2O production mechanisms were complex and could involve multiple N2O production pathways. (C) 2013 Elsevier Ltd. All rights reserved.
  • 押木 守, 佐藤 久, 岡部 聡 日本生物工学会大会講演要旨集 65 2013年08月25日 [査読無し][通常論文]
  • 羽深昭, 吉川弘晃, 山田幸司, 高橋正宏, 岡部聡, 佐藤久 分析化学討論会講演要旨集 73rd 149 2013年05月04日 [査読無し][通常論文]
  • W. M. K. R. T. W. Bandara, M. Ikeda, H. Satoh, M. Sasakawa, Y. Nakahara, M. Takahashi, S. Okabe WATER ENVIRONMENT RESEARCH 85 (5) 387 -390 2013年05月 [査読無し][通常論文]
     
    The effectiveness of degasification using a degassing membrane to improve chemical oxygen demand (COD) removal efficiency was investigated using a bench-scale upflow anaerobic sludge blanket (UASB) reactor. Vacuum degasification was able to transfer dissolved gas in the bulk liquid of the UASB reactor inside the membrane. Such a process might provide thermodynamically favorable conditions for the degradation of organic compounds. The COD-removal efficiency improved from 83% during normal operation to 90% during the degassing operation.
  • 井川裕介, 吉田圭太郎, 福島寿和, 岡部聡, 三好太郎, 渡辺義公 日本水環境学会年会講演集 47th 255 2013年03月11日 [査読無し][通常論文]
  • 吉川弘晃, 羽深昭, 菅藤亮輔, 大屋光平, 山田幸司, 高橋正宏, 岡部聡, 佐藤久 日本水環境学会年会講演集 47th 431 2013年03月11日 [査読無し][通常論文]
  • 宮崎悠爾, 谷内翔, 高橋正宏, 岡部聡, 佐藤久 日本水環境学会年会講演集 47th 321 2013年03月11日 [査読無し][通常論文]
  • 菅藤亮輔, 羽深昭, 吉川弘晃, 大屋光平, 山田幸司, 高橋正宏, 岡部聡, 佐藤久 日本水環境学会年会講演集 47th 432 2013年03月11日 [査読無し][通常論文]
  • 大屋光平, 羽深昭, 菅藤亮輔, 吉川弘晃, 山田幸司, 高橋正宏, 岡部聡, 佐藤久 日本水環境学会年会講演集 47th 692 2013年03月11日 [査読無し][通常論文]
  • 谷内翔, 宮崎悠爾, 高橋正宏, 岡部聡, 佐藤久 日本水環境学会年会講演集 47th 322 2013年03月11日 [査読無し][通常論文]
  • 坂槙有紀恵, 山田健太, 佐野大輔, 高橋正宏, 岡部聡, 佐藤久 日本水環境学会年会講演集 47th 436 2013年03月11日 [査読無し][通常論文]
  • 中島弘司, 原(山村)宏江, 木村克輝, 岡部聡, 三好太郎, 渡辺義公 日本水環境学会年会講演集 47th 412 2013年03月11日 [査読無し][通常論文]
  • 福島寿和, 木村克輝, 岡部聡, 三好太郎, 渡辺義公 日本水環境学会年会講演集 47th 516 2013年03月11日 [査読無し][通常論文]
  • 原(山村)宏江, HOQUE Asiful, 木村克輝, 岡部聡, 三好太郎, 渡辺義公 日本水環境学会年会講演集 47th 517 2013年03月11日 [査読無し][通常論文]
  • 粟田貴宣, 金田一智規, 尾崎則篤, 大橋晶良, 押木守, 岡部聡 日本水環境学会年会講演集 47th 160 2013年03月11日 [査読無し][通常論文]
  • 宮崎悠爾, 谷内翔, 押木守, 佐藤久, 高橋正宏, 岡部聡 土木学会北海道支部論文報告集(CD-ROM) (69) ROMBUNNO.G-02 2013年02月 [査読無し][通常論文]
  • 宮崎悠爾, 谷内翔, 押木守, 高橋正宏, 岡部聡, 佐藤久 衛生工学シンポジウム論文集 20th 24 2012年12月04日 [査読無し][通常論文]
  • 坂槙有紀恵, 山田健太, ピタックティーラタム ニティ, 石井聡, 佐野大輔, 高橋正宏, 岡部聡, 佐藤久 環境工学研究フォーラム講演集 49th 28 -30 2012年11月28日 [査読無し][通常論文]
  • 菅藤亮輔, 羽深昭, 吉川弘晃, 大屋光平, 山田幸司, 高橋正宏, 岡部聡, 佐藤久 環境工学研究フォーラム講演集 49th 31 -33 2012年11月28日 [査読無し][通常論文]
  • 宮崎悠爾, 谷内翔, 押木守, 佐藤久, 高橋正宏, 岡部聡 環境工学研究フォーラム講演集 49th 25 -27 2012年11月28日 [査読無し][通常論文]
  • 木村 善一郎, 寺田 浩太朗, 岡部 聡 日本微生物生態学会講演要旨集 (28) 2012年09月19日 [査読無し][通常論文]
  • 押木 守, 新家子 香織, 佐藤 久, 岡部 聡 日本微生物生態学会講演要旨集 (28) 2012年09月19日 [査読無し][通常論文]
  • 佐藤久, 坂槙有紀恵, 山田健太, ピタックティーラタムニティ, 石井聡, 佐野大輔, 高橋正宏, 岡部聡 日本水環境学会シンポジウム講演集 15th 233 -234 2012年09月10日 [査読無し][通常論文]
  • 押木守, 佐藤久, 岡部聡, 新家子香織 日本水環境学会シンポジウム講演集 15th 157 2012年09月10日 [査読無し][通常論文]
  • 谷内翔, 宮崎悠爾, 高橋正宏, 岡部聡, 佐藤久 日本分析化学会年会講演要旨集 61st 324 2012年09月05日 [査読無し][通常論文]
  • 吉川弘晃, 菅藤亮輔, 羽深昭, 山田幸司, 高橋正宏, 岡部聡, 佐藤久 日本分析化学会年会講演要旨集 61st 315 2012年09月05日 [査読無し][通常論文]
  • 坂槙有紀恵, 山田健太, 高橋正宏, 岡部聡, 佐藤久 日本分析化学会年会講演要旨集 61st 397 2012年09月05日 [査読無し][通常論文]
  • バイオフィルム感染症の新治療戦略 大腸菌RelEが誘導する菌体密度依存的休眠化
    岡部 聡, 田代 陽介, 谷内 亜沙美, Thithiwat May, 柿沼 建至, 川田 耕司 日本生物工学会大会講演要旨集 平成24年度 100 -100 2012年09月 [査読無し][通常論文]
  • 菅藤亮輔, 吉川弘晃, 谷山拓生, 羽深昭, 山田幸司, 高橋正宏, 岡部聡, 佐藤久 分析化学討論会講演要旨集 72nd 130 2012年05月05日 [査読無し][通常論文]
  • 日向寺崇文, 前田高輝, 岡部聡, 高橋正宏, 佐藤久 日本水環境学会年会講演集 46th 510 2012年03月14日 [査読無し][通常論文]
  • 谷山拓生, 羽深昭, 菅藤亮輔, 吉川弘晃, 山田幸司, 高橋正宏, 岡部聡, 佐藤久 日本水環境学会年会講演集 46th 506 2012年03月14日 [査読無し][通常論文]
  • 吉川弘晃, 羽深昭, 谷山拓生, 菅藤亮輔, 山田幸司, 高橋正宏, 岡部聡, 佐藤久 日本水環境学会年会講演集 46th 676 2012年03月14日 [査読無し][通常論文]
  • 羽深昭, 谷山拓生, 菅藤亮輔, 吉川弘晃, 山田幸司, 高橋正宏, 岡部聡, 佐藤久 日本水環境学会年会講演集 46th 622 2012年03月14日 [査読無し][通常論文]
  • 中島弘司, 原宏江, HOQUE Asiful, 木村克輝, 岡部聡, 三好太郎, 渡辺義公 日本水環境学会年会講演集 46th 93 2012年03月14日 [査読無し][通常論文]
  • 原(山村)宏江, HOQUE Asiful, 木村克輝, 岡部聡, 中島弘司, 三好太郎, 渡辺義公 日本水環境学会年会講演集 46th 74 2012年03月14日 [査読無し][通常論文]
  • 石黒真規, 押木守, 石井聡, 岡部聡 日本水環境学会年会講演集 46th 151 2012年03月14日 [査読無し][通常論文]
  • 平泉晴菜, 押木守, 岡部聡 日本水環境学会年会講演集 46th 152 2012年03月14日 [査読無し][通常論文]
  • 宮崎悠爾, 谷内翔, 高橋正宏, 岡部聡, 佐藤久 化学系学協会北海道支部冬季研究発表会講演要旨集 2012 148 2012年01月31日 [査読無し][通常論文]
  • 菅藤亮輔, 吉川弘晃, 谷山拓生, 羽深昭, 山田幸司, 高橋正宏, 岡部聡, 佐藤久 化学系学協会北海道支部冬季研究発表会講演要旨集 2012 101 2012年01月31日 [査読無し][通常論文]
  • H. Satoh, I. Tsushima, Y. Miura, T. Ito, S. Okabe WATER SCIENCE AND TECHNOLOGY 65 (12) 2125 -2131 2012年 [査読無し][通常論文]
     
    The spatial distribution of microorganisms and their in situ activities in anaerobic granules were investigated by fluorescence in situ hybridization (FISH), beta imaging and microsensors. FISH results revealed a layered structure of microorganisms in the granule, where Chloroflexi was present in the outermost layer, Smithella spp. and Syntrophobacter spp. were found in a depth of ca. 100 mu m, and Archaea was restricted to the inner layer (below ca. 300 mu m from the surface). Substrate uptake patterns elucidated by beta imaging demonstrated that glucose uptake was highest at 50 mu m depth, whereas propionate uptake had a peak at 200 mu m depth. In addition, microsensor measurements revealed that acid was produced mainly at 100 mu m depth and H-2 production was detected at a depth from 100 to 200 mu m. H-2 consumption and corresponding CH4 production were found below 200 mu m from the surface. Direct comparison of these results implied sequential degradation of complex organic compounds in anaerobic granules; Chloroflexi contributed to fermentation of organic compounds and acid production in the outermost layer, volatile fatty acids were oxidized and H-2 was produced mainly by Smithella spp. and Syntrophobacter spp. at a depth from 100 to 200 mu m, and Archaea produced CH4 below ca. 300 mu m from the surface.
  • Takahiro Imai, Daisuke Sano, Takayuki Miura, Satoshi Okabe, Keishi Wada, Yoshifumi Masago, Tatsuo Omura BMC BIOTECHNOLOGY 11 (123) 1 -11 2011年12月 [査読無し][通常論文]
     
    Background: Water contamination with human enteric viruses has posed human health risks all over the world. Reasonable and facile methodologies for recovering and quantifying infectious enteric viruses in environmental samples are needed to address the issues of waterborne viral infectious diseases. In this study, a bacterial protein that has a binding capability with several enteric viruses is discovered, and its binding characteristics were investigated for utilizing it as a viral adsorbent in virus recovery and detection technologies. Results: A gene of an enteric virus-binding protein (EVBP), derived from a monomer of a bacterial chaperon protein GroEL, was successfully acquired from a genomic DNA library of activated sludge microorganisms with nested PCR. Equilibrium dissociation constants between EVBP and norovirus-like particles (NoVLPs) of genotypes GI.7 and GII.4, estimated with quartz crystal microbalance method, were 240 and 210 nM, respectively. These values of equilibrium dissociation constant imply that the binding affinity between EVBP and NoVLPs is 1 to 3-log weaker than that in general antigen-antibody interactions, but about 2-log stronger than that in weak specific interactions of proteins with cations and organic polymers. The adsorptions of EVBP to norovirus, group A rotavirus and poliovirus type 1 were found to be significant in enzyme-linked immunosorbent assay. Meanwhile, the binding of native GroEL tetradecamer to viral particles was weaker than that of EVBP, presumably because of a steric hindrance. The small molecule of EVBP could have an advantage in the access to the surface of viral particles with rugged structure. Conclusions: EVBP that has a broad binding spectrum to enteric viruses was newly discovered. The broad binding characteristic of EVBP would allow us to utilize it as a novel adsorbent for detecting diverse enteric viruses in clinical and environmental samples.
  • Thithiwat May, Satoshi Okabe ENVIRONMENTAL MICROBIOLOGY 13 (12) 3149 -3162 2011年12月 [査読無し][通常論文]
     
    A variety of bacterial cell surface structures and quorum signalling molecules play a role in biofilm development in Escherichia coli. However, here we show that an engineered reduced-genome E. coli mutant that lacks 17.6% of the parental E. coli genome, including the genes involved in the synthesis of various cell surface structures, such as type 1 fimbriae, curli, exopolysaccharide polymers and the autoinducer-2 signalling molecule, is able to develop mature biofilms. Using temporal gene expression profiling, we investigated phenotypic changes in reduced-genome biofilms in relation with the genes encoding the synthesis of different amino acids that were differentially expressed during biofilm formation. We identified and characterized entB, marR, dosC, mcbR and yahK genes, as involved in biofilm formation by the reduced-genome E. coli. Of these, for a first time, we demonstrated that overproduction of entB and yahK, which encode an enterobactin for iron transport and a hypothetical oxidoreductase protein, respectively, promoted biofilm development and maturation. Our results indicate that specific types of genes contribute to phenotypic changes in reduced-genome E. coli biofilms. In addition, this work demonstrates that the functions of biofilm-specific genes could be analysed through experiments using the reduced-genome E. coli.
  • OKABE Satoshi, OKABE Satoshi, OSHIKI Mamoru, OSHIKI Mamoru, TAKAHASHI Yoshitaka, SATOH Hisashi Water Research 45 (19) 6461 -6470 2011年12月 [査読無し][通常論文]
     
    Emission of nitrous oxide (N2O) during biological wastewater treatment is of growing concern. The emission of N2O from a lab-scale two-reactor partial nitrification (PN)-anammox reactor was therefore determined in this study. The average emission of N2O from the PN and anammox process was 4.0 +/- 1.5% (9.6 +/- 3.2% of the removed nitrogen) and 0.1 +/- 0.07% (0.14 +/- 0.09% of the removed nitrogen) of the incoming nitrogen load, respectively. Thus, a larger part (97.5%) of N2O was emitted from the PN reactor. The total amount of N2O emission from the PN reactor was correlated to nitrite (NO2-) concentration in the PN effluent rather than DO concentration. In addition, further studies were performed to indentify a key biological process that is responsible for N2O emission from the anammox process (i.e., granules). In order to characterize N2O emission from the anammox granules, the in situ N2O production rate was determined by using microelectrodes for the first time, which was related to the spatial organization of microbial community of the granule as determined by fluorescence in situ hybridization (FISH). Microelectrode measurement revealed that the active N2O production zone was located in the inner part of the anammox granule, whereas the active ammonium consumption zone was located above the N2O production zone. Anammox bacteria were present throughout the granule, whereas ammonium-oxidizing bacteria (AOB) were restricted to only the granule surface. In addition, addition of penicillin G that inhibits most of the heterotrophic denitrifiers and AOB completely inhibited N2O production in batch experiments. Based on these results obtained, denitrification by putative heterotrophic denitrifiers present in the inner part of the granule was considered the most probable cause of N2O emission from the anammox reactor (i.e., granules). (C) 2011 Elsevier Ltd. All rights reserved.
  • Tsukasa Ito, Kazumi Yoshiguchi, Herto Dwi Ariesyady, Satoshi Okabe ISME JOURNAL 5 (12) 1844 -1856 2011年12月 [査読無し][通常論文]
     
    Major acetate-utilizing bacterial and archaeal populations in methanogenic anaerobic digester sludge were identified and quantified by radioisotope-and stable-isotope-based functional analyses, microautoradiography-fluorescence in situ hybridization (MAR-FISH) and stable-isotope probing of 16S rRNA (RNA-SIP) that can directly link 16S rRNA phylogeny with in situ metabolic function. First, MAR-FISH with (14)C-acetate indicated the significant utilization of acetate by only two major groups, unidentified bacterial cells and Methanosaeta-like filamentous archaeal cells, in the digester sludge. To identify the acetate-utilizing unidentified bacteria, RNA-SIP was conducted with (13)C(6)-glucose and (13)C(3)-propionate as sole carbon source, which were followed by phylogenetic analysis of 16S rRNA. We found that bacteria belonging to Synergistes group 4 were commonly detected in both 16S rRNA clone libraries derived from the sludge incubated with (13)C-glucose and (13)C-propionate. To confirm that this bacterial group can utilize acetate, specific FISH probe targeting for Synergistes group 4 was newly designed and applied to the sludge incubated with (14)C-acetate for MAR-FISH. The MAR-FISH result showed that bacteria belonging to Synergistes group 4 significantly took up acetate and their active population size was comparable to that of Methanosaeta in this sludge. In addition, as bacteria belonging to Synergistes group 4 had high K(m) for acetate and maximum utilization rate, they are more competitive for acetate over Methanosaeta at high acetate concentrations (2.5-10mM). To our knowledge, it is the first time to report the acetate-utilizing activity of uncultured bacteria belonging to Synergistes group 4 and its competitive significance to acetoclastic methanogen, Methanosaeta.The ISME Journal (2011) 5, 1844-1856; doi:10.1038/ismej.2011.59; published online 12 May 2011
  • Takahiro Imai, Daisuke Sano, Takayuki Miura, Satoshi Okabe, Keishi Wada, Yoshifumi Masago, Tatsuo Omura BMC BIOTECHNOLOGY 11 (123) 1 -11 2011年12月 [査読無し][通常論文]
     
    Background: Water contamination with human enteric viruses has posed human health risks all over the world. Reasonable and facile methodologies for recovering and quantifying infectious enteric viruses in environmental samples are needed to address the issues of waterborne viral infectious diseases. In this study, a bacterial protein that has a binding capability with several enteric viruses is discovered, and its binding characteristics were investigated for utilizing it as a viral adsorbent in virus recovery and detection technologies. Results: A gene of an enteric virus-binding protein (EVBP), derived from a monomer of a bacterial chaperon protein GroEL, was successfully acquired from a genomic DNA library of activated sludge microorganisms with nested PCR. Equilibrium dissociation constants between EVBP and norovirus-like particles (NoVLPs) of genotypes GI.7 and GII.4, estimated with quartz crystal microbalance method, were 240 and 210 nM, respectively. These values of equilibrium dissociation constant imply that the binding affinity between EVBP and NoVLPs is 1 to 3-log weaker than that in general antigen-antibody interactions, but about 2-log stronger than that in weak specific interactions of proteins with cations and organic polymers. The adsorptions of EVBP to norovirus, group A rotavirus and poliovirus type 1 were found to be significant in enzyme-linked immunosorbent assay. Meanwhile, the binding of native GroEL tetradecamer to viral particles was weaker than that of EVBP, presumably because of a steric hindrance. The small molecule of EVBP could have an advantage in the access to the surface of viral particles with rugged structure. Conclusions: EVBP that has a broad binding spectrum to enteric viruses was newly discovered. The broad binding characteristic of EVBP would allow us to utilize it as a novel adsorbent for detecting diverse enteric viruses in clinical and environmental samples.
  • Thithiwat May, Satoshi Okabe ENVIRONMENTAL MICROBIOLOGY 13 (12) 3149 -3162 2011年12月 [査読無し][通常論文]
     
    A variety of bacterial cell surface structures and quorum signalling molecules play a role in biofilm development in Escherichia coli. However, here we show that an engineered reduced-genome E. coli mutant that lacks 17.6% of the parental E. coli genome, including the genes involved in the synthesis of various cell surface structures, such as type 1 fimbriae, curli, exopolysaccharide polymers and the autoinducer-2 signalling molecule, is able to develop mature biofilms. Using temporal gene expression profiling, we investigated phenotypic changes in reduced-genome biofilms in relation with the genes encoding the synthesis of different amino acids that were differentially expressed during biofilm formation. We identified and characterized entB, marR, dosC, mcbR and yahK genes, as involved in biofilm formation by the reduced-genome E. coli. Of these, for a first time, we demonstrated that overproduction of entB and yahK, which encode an enterobactin for iron transport and a hypothetical oxidoreductase protein, respectively, promoted biofilm development and maturation. Our results indicate that specific types of genes contribute to phenotypic changes in reduced-genome E. coli biofilms. In addition, this work demonstrates that the functions of biofilm-specific genes could be analysed through experiments using the reduced-genome E. coli.
  • Tsukasa Ito, Kazumi Yoshiguchi, Herto Dwi Ariesyady, Satoshi Okabe ISME JOURNAL 5 (12) 1844 -1856 2011年12月 [査読無し][通常論文]
     
    Major acetate-utilizing bacterial and archaeal populations in methanogenic anaerobic digester sludge were identified and quantified by radioisotope-and stable-isotope-based functional analyses, microautoradiography-fluorescence in situ hybridization (MAR-FISH) and stable-isotope probing of 16S rRNA (RNA-SIP) that can directly link 16S rRNA phylogeny with in situ metabolic function. First, MAR-FISH with (14)C-acetate indicated the significant utilization of acetate by only two major groups, unidentified bacterial cells and Methanosaeta-like filamentous archaeal cells, in the digester sludge. To identify the acetate-utilizing unidentified bacteria, RNA-SIP was conducted with (13)C(6)-glucose and (13)C(3)-propionate as sole carbon source, which were followed by phylogenetic analysis of 16S rRNA. We found that bacteria belonging to Synergistes group 4 were commonly detected in both 16S rRNA clone libraries derived from the sludge incubated with (13)C-glucose and (13)C-propionate. To confirm that this bacterial group can utilize acetate, specific FISH probe targeting for Synergistes group 4 was newly designed and applied to the sludge incubated with (14)C-acetate for MAR-FISH. The MAR-FISH result showed that bacteria belonging to Synergistes group 4 significantly took up acetate and their active population size was comparable to that of Methanosaeta in this sludge. In addition, as bacteria belonging to Synergistes group 4 had high K(m) for acetate and maximum utilization rate, they are more competitive for acetate over Methanosaeta at high acetate concentrations (2.5-10mM). To our knowledge, it is the first time to report the acetate-utilizing activity of uncultured bacteria belonging to Synergistes group 4 and its competitive significance to acetoclastic methanogen, Methanosaeta.The ISME Journal (2011) 5, 1844-1856; doi:10.1038/ismej.2011.59; published online 12 May 2011
  • 羽深昭, 谷山拓生, 菅藤亮輔, 吉川弘晃, 山田幸司, 高橋正宏, 岡部聡, 佐藤久 環境工学研究フォーラム講演集 48th 214 -216 2011年11月25日 [査読無し][通常論文]
  • 菅藤亮輔, 羽深昭, 谷山拓生, 吉川弘晃, 山田幸司, 高橋正宏, 岡部聡, 佐藤久 環境工学研究フォーラム講演集 48th 220 -222 2011年11月25日 [査読無し][通常論文]
  • 谷内翔, 宮崎悠爾, 押木守, 佐藤久, 高橋正宏, 岡部聡 環境工学研究フォーラム講演集 48th 217 -219 2011年11月25日 [査読無し][通常論文]
  • 木村善一郎, 伊藤皓亮, 岡部聡 日本微生物生態学会講演要旨集 27th 74 2011年10月08日 [査読無し][通常論文]
  • 平泉 晴菜, 新家子 香織, 押木 守, 佐藤 久, 岡部 聡 日本微生物生態学会講演要旨集 (27) 2011年10月08日 [査読無し][通常論文]
  • 押木 守, 新家子 香織, 佐藤 久, 岡部 聡 日本微生物生態学会講演要旨集 (27) 2011年10月08日 [査読無し][通常論文]
  • 岡部聡, 押木守, 高橋慶考, 佐藤久 日本水環境学会シンポジウム講演集 14th 43 -44 2011年09月10日 [査読無し][通常論文]
  • 谷山拓生, 羽深昭, 山田幸司, 高橋正宏, 岡部聡, 佐藤久 土木学会年次学術講演会講演概要集(CD-ROM) 66th ROMBUNNO.VII-192 2011年08月05日 [査読無し][通常論文]
  • 羽深昭, 谷山拓生, 山田幸司, 高橋正宏, 岡部聡, 佐藤久 土木学会年次学術講演会講演概要集(CD-ROM) 66th ROMBUNNO.VII-191 2011年08月05日 [査読無し][通常論文]
  • 宮崎悠爾, 押木守, 高橋正宏, 岡部聡, 佐藤久 土木学会年次学術講演会講演概要集(CD-ROM) 66th ROMBUNNO.VII-029 2011年08月05日 [査読無し][通常論文]
  • 谷山拓生, 菅藤亮輔, 羽深昭, 山田幸司, 高橋正宏, 岡部聡, 佐藤久 日本化学会北海道支部夏季研究発表会講演要旨集 2011 150 2011年07月01日 [査読無し][通常論文]
  • 菅藤亮輔, 谷山拓生, 羽深昭, 山田幸司, 高橋正宏, 岡部聡, 佐藤久 日本化学会北海道支部夏季研究発表会講演要旨集 2011 149 2011年07月01日 [査読無し][通常論文]
  • Satoshi Okabe, Mamoru Oshiki, Yoshitaka Takahashi, Flisashi Satoh BIORESOURCE TECHNOLOGY 102 (13) 6801 -6807 2011年07月 [査読無し][通常論文]
     
    The partial nitrification reactor was successfully started up and operated stably for more than 250 days with a maximum nitrite production rate of 1.12 kg-N m(-3) day(-1). The important factors for successful partial nitrification were high ammonium loading rate (>1.0 kg-N m(-3) day(-1)) and relatively high pH (ca. 8.0), giving high free ammonia concentrations (>10 mg NH(3)-N L(-1)). In addition, the air flow rate must be controlled at the ratio of air flow rate to ammonium loading rate below 0.1 (m(air)(3) day(-1))/(kg-N m(-3) day(-1)). After the establishment of stable partial nitrification, the effluent NO(2)(-)-N/NH(4)(+)-N ratio and effluent NO(3)(-)-N concentration were 1.20 +/- 0.33 and 1.2 +/- 1.0 mg-N L(-1), respectively, which was then fed into an granular-sludge anammox reactor. Consistent nitrogen removal was achieved for more than 250 days with a maximum nitrogen removal rate of 15.0 kg-TN m(-3) day(-1). (C) 2011 Elsevier Ltd. All rights reserved.
  • Satoshi Okabe, Mamoru Oshiki, Yoshitaka Takahashi, Flisashi Satoh BIORESOURCE TECHNOLOGY 102 (13) 6801 -6807 2011年07月 [査読無し][通常論文]
     
    The partial nitrification reactor was successfully started up and operated stably for more than 250 days with a maximum nitrite production rate of 1.12 kg-N m(-3) day(-1). The important factors for successful partial nitrification were high ammonium loading rate (>1.0 kg-N m(-3) day(-1)) and relatively high pH (ca. 8.0), giving high free ammonia concentrations (>10 mg NH(3)-N L(-1)). In addition, the air flow rate must be controlled at the ratio of air flow rate to ammonium loading rate below 0.1 (m(air)(3) day(-1))/(kg-N m(-3) day(-1)). After the establishment of stable partial nitrification, the effluent NO(2)(-)-N/NH(4)(+)-N ratio and effluent NO(3)(-)-N concentration were 1.20 +/- 0.33 and 1.2 +/- 1.0 mg-N L(-1), respectively, which was then fed into an granular-sludge anammox reactor. Consistent nitrogen removal was achieved for more than 250 days with a maximum nitrogen removal rate of 15.0 kg-TN m(-3) day(-1). (C) 2011 Elsevier Ltd. All rights reserved.
  • 木村善一郎, 伊藤皓亮, 岡部聡 環境バイオテクノロジー学会大会プログラム講演要旨集 44th 25 2011年06月20日 [査読無し][通常論文]
  • Mamoru Oshiki, Masaki Shimokawa, Naoki Fujii, Hisashi Satohl, Satoshi Okabe MICROBIOLOGY-SGM 157 (6) 1706 -1713 2011年06月 [査読無し][通常論文]
     
    The present study investigated the phylogenetic affiliation and physiological characteristics of bacteria responsible for anaerobic ammonium oxidization (anammox); these bacteria were enriched in an anammox reactor with a nitrogen removal rate of 26.0 kg N m(-3) day(-1). The anammox bacteria were identified as representing 'Candidatus Brocadia sinica' on the basis of phylogenetic analysis of rRNA operon sequences. Physiological characteristics examined were growth rate, kinetics of ammonium oxidation and nitrite reduction, temperature, pH and inhibition of anammox. The maximum specific growth rate (mu(max).) was 0.0041 h(-1), corresponding to a doubling time of 7 days. The half-saturation constants (K(s)) for ammonium and nitrite of 'Ca. B. sinica' were 28 +/- 4 and 86 +/- 4 mu M, respectively, higher than those of 'Candidatus Brocadia anammoxidans' and 'Candidatus Kuenenia stuttgartiensis'. The temperature and pH ranges of anammox activity were 25-45 degrees C and pH 6.5-8.8, respectively. Anammox activity was inhibited in the presence of nitrite (50% inhibition at 16 mM), ethanol (91% at 1 mM) and methanol (86% at 1 mM). Anammox activities were 80 and 70% of baseline in the presence of 20 mM phosphorus and 3% salinity, respectively. The yield of biomass and dissolved organic carbon production in the culture supernatant were 0.062 and 0.005 mol C (mol NH(4)(+))(-1), respectively. This study compared physiological differences between three anammox bacterial enrichment cultures to provide a better understanding of anammox niche specificity in natural and man-made ecosystems.
  • Mamoru Oshiki, Masaki Shimokawa, Naoki Fujii, Hisashi Satohl, Satoshi Okabe MICROBIOLOGY-SGM 157 (6) 1706 -1713 2011年06月 [査読無し][通常論文]
     
    The present study investigated the phylogenetic affiliation and physiological characteristics of bacteria responsible for anaerobic ammonium oxidization (anammox); these bacteria were enriched in an anammox reactor with a nitrogen removal rate of 26.0 kg N m(-3) day(-1). The anammox bacteria were identified as representing 'Candidatus Brocadia sinica' on the basis of phylogenetic analysis of rRNA operon sequences. Physiological characteristics examined were growth rate, kinetics of ammonium oxidation and nitrite reduction, temperature, pH and inhibition of anammox. The maximum specific growth rate (mu(max).) was 0.0041 h(-1), corresponding to a doubling time of 7 days. The half-saturation constants (K(s)) for ammonium and nitrite of 'Ca. B. sinica' were 28 +/- 4 and 86 +/- 4 mu M, respectively, higher than those of 'Candidatus Brocadia anammoxidans' and 'Candidatus Kuenenia stuttgartiensis'. The temperature and pH ranges of anammox activity were 25-45 degrees C and pH 6.5-8.8, respectively. Anammox activity was inhibited in the presence of nitrite (50% inhibition at 16 mM), ethanol (91% at 1 mM) and methanol (86% at 1 mM). Anammox activities were 80 and 70% of baseline in the presence of 20 mM phosphorus and 3% salinity, respectively. The yield of biomass and dissolved organic carbon production in the culture supernatant were 0.062 and 0.005 mol C (mol NH(4)(+))(-1), respectively. This study compared physiological differences between three anammox bacterial enrichment cultures to provide a better understanding of anammox niche specificity in natural and man-made ecosystems.
  • Wasala M. K. R. T. W. Bandara, Hisashi Satoh, Manabu Sasakawa, Yoshihito Nakahara, Masahiro Takahashi, Satoshi Okabe WATER RESEARCH 45 (11) 3533 -3540 2011年05月 [査読無し][通常論文]
     
    In this study, we investigated the efficiency of dissolved methane (D-CH(4)) collection by degasification from the effluent of a bench-scale upflow anaerobic sludge blanket (UASB) reactor treating synthetic wastewater. A hollow-fiber degassing membrane module was used for degasification. This module was connected to the liquid outlet of the UASB reactor. After chemical oxygen demand (COD) removal efficiency of the UASB reactor became stable, D-CH(4) discharged from the UASB reactor was collected. Under 35 degrees C and a hydraulic retention time (HRT) of 10 h, average D-CH(4) concentration could be reduced from 63 mg COD L(-1) to 15 mg COD L(-1); this, in turn, resulted in an increase in total methane (CH(4)) recovery efficiency from 89% to 97%. Furthermore, we investigated the effects of temperature and HRT of the UASB reactor on degasification efficiency. Average D-CH(4) concentration was as high as 104 mg COD L(-1) at 15 degrees C because of the higher solubility of CH(4) gas in liquid; the average D-CH(4) concentration was reduced to 14 mg COD L(-1) by degasification. Accordingly, total CH(4) recovery efficiency increased from 71% to 97% at 15 degrees C as a result of degasification. Moreover, degasification tended to cause an increase in particulate COD removal efficiency. The UASB reactor was operated at the same COD loading rate, but different wastewater feed rates and HRTs. Although average D-CH(4) concentration in the UASB reactor was almost unchanged (ca. 70 mg COD L(-1)) regardless of the HRT value, the CH(4) discharge rate from the UASB reactor increased because of an increase in the wastewater feed rate. Because the D-CH(4) concentration could be reduced down to 12 +/- 1 mg COD L(-1) by degasification at an HRT of 6.7 h, the CH(4) recovery rate was 1.5 times higher under degasification than under normal operation. (C) 2011 Elsevier Ltd. All rights reserved.
  • Wasala M. K. R. T. W. Bandara, Hisashi Satoh, Manabu Sasakawa, Yoshihito Nakahara, Masahiro Takahashi, Satoshi Okabe WATER RESEARCH 45 (11) 3533 -3540 2011年05月 [査読無し][通常論文]
     
    In this study, we investigated the efficiency of dissolved methane (D-CH(4)) collection by degasification from the effluent of a bench-scale upflow anaerobic sludge blanket (UASB) reactor treating synthetic wastewater. A hollow-fiber degassing membrane module was used for degasification. This module was connected to the liquid outlet of the UASB reactor. After chemical oxygen demand (COD) removal efficiency of the UASB reactor became stable, D-CH(4) discharged from the UASB reactor was collected. Under 35 degrees C and a hydraulic retention time (HRT) of 10 h, average D-CH(4) concentration could be reduced from 63 mg COD L(-1) to 15 mg COD L(-1); this, in turn, resulted in an increase in total methane (CH(4)) recovery efficiency from 89% to 97%. Furthermore, we investigated the effects of temperature and HRT of the UASB reactor on degasification efficiency. Average D-CH(4) concentration was as high as 104 mg COD L(-1) at 15 degrees C because of the higher solubility of CH(4) gas in liquid; the average D-CH(4) concentration was reduced to 14 mg COD L(-1) by degasification. Accordingly, total CH(4) recovery efficiency increased from 71% to 97% at 15 degrees C as a result of degasification. Moreover, degasification tended to cause an increase in particulate COD removal efficiency. The UASB reactor was operated at the same COD loading rate, but different wastewater feed rates and HRTs. Although average D-CH(4) concentration in the UASB reactor was almost unchanged (ca. 70 mg COD L(-1)) regardless of the HRT value, the CH(4) discharge rate from the UASB reactor increased because of an increase in the wastewater feed rate. Because the D-CH(4) concentration could be reduced down to 12 +/- 1 mg COD L(-1) by degasification at an HRT of 6.7 h, the CH(4) recovery rate was 1.5 times higher under degasification than under normal operation. (C) 2011 Elsevier Ltd. All rights reserved.
  • Thithiwat May, Kenji Tsuruta, Satoshi Okabe ISME JOURNAL 5 (4) 771 -775 2011年04月 [査読無し][通常論文]
     
    Escherichia coli carrying a natural conjugative F-plasmid generates F-pili mating pairs, which is important for early biofilm formation. In this study, we investigated the effect of male-specific filamentous single stranded DNA bacteriophage (f1) and RNA bacteriophage (MS2) on the formation of biofilms by E. coli carrying a natural conjugative F-plasmid. We showed that the early biofilm formation was completely inhibited by addition of the f1 phage, but not the MS2 phage. This suggests that the tip of F-pili is the specific attachment site for mating pairs formation and the side of F-pili has a non-obligatory role during biofilm formation. The inhibitory effect of the f1 phage was dependent on the time of addition during the biofilm formation. No inhibitory effect was observed when the f1 phages were added to the mature biofilms. This resistant mechanism of the mature biofilms could be attributed to the biofilm-specific phenotypes representing that the F-pili mating pairs were already formed and then the curli production commenced during the biofilm maturation. The pre-formed mating pairs seemed to resist the f1 phages. Altogether, our results indicate a close relationship between the presence of conjugative plasmid and male-specific bacteriophages within sessile biofilm communities, as well as the possibility of using the male-specific bacteriophages to control biofilm formation. The ISME Journal (2011) 5, 771-775; doi: 10.1038/ismej.2010.158; published online 21 October 2010
  • Thithiwat May, Kenji Tsuruta, Satoshi Okabe ISME JOURNAL 5 (4) 771 -775 2011年04月 [査読無し][通常論文]
     
    Escherichia coli carrying a natural conjugative F-plasmid generates F-pili mating pairs, which is important for early biofilm formation. In this study, we investigated the effect of male-specific filamentous single stranded DNA bacteriophage (f1) and RNA bacteriophage (MS2) on the formation of biofilms by E. coli carrying a natural conjugative F-plasmid. We showed that the early biofilm formation was completely inhibited by addition of the f1 phage, but not the MS2 phage. This suggests that the tip of F-pili is the specific attachment site for mating pairs formation and the side of F-pili has a non-obligatory role during biofilm formation. The inhibitory effect of the f1 phage was dependent on the time of addition during the biofilm formation. No inhibitory effect was observed when the f1 phages were added to the mature biofilms. This resistant mechanism of the mature biofilms could be attributed to the biofilm-specific phenotypes representing that the F-pili mating pairs were already formed and then the curli production commenced during the biofilm maturation. The pre-formed mating pairs seemed to resist the f1 phages. Altogether, our results indicate a close relationship between the presence of conjugative plasmid and male-specific bacteriophages within sessile biofilm communities, as well as the possibility of using the male-specific bacteriophages to control biofilm formation. The ISME Journal (2011) 5, 771-775; doi: 10.1038/ismej.2010.158; published online 21 October 2010
  • 藤木一到, 石崎創, 木村善一郎, 岡部聡 日本水環境学会年会講演集 45th 168 2011年03月18日 [査読無し][通常論文]
  • 木村善一郎, 岡部聡 日本水環境学会年会講演集 45th 166 2011年03月18日 [査読無し][通常論文]
  • 谷山拓生, 羽深昭, 菅藤亮輔, 山田幸司, 高橋正宏, 岡部聡, 佐藤久 日本水環境学会年会講演集 45th 293 2011年03月18日 [査読無し][通常論文]
  • 菅藤亮輔, 羽深昭, 谷山拓生, 山田幸司, 高橋正宏, 岡部聡, 佐藤久 日本水環境学会年会講演集 45th 295 2011年03月18日 [査読無し][通常論文]
  • 羽深昭, 谷山拓生, 菅藤亮輔, 山田幸司, 高橋正宏, 岡部聡, 佐藤久 日本水環境学会年会講演集 45th 294 2011年03月18日 [査読無し][通常論文]
  • SONG Yanjun, 押木守, 岡部聡 日本水環境学会年会講演集 45th 161 2011年03月18日 [査読無し][通常論文]
  • 平泉晴菜, 新家子香織, 押木守, 岡部聡 日本水環境学会年会講演集 45th 47 2011年03月18日 [査読無し][通常論文]
  • 新家子香織, 押木守, 佐藤久, 岡部聡 日本水環境学会年会講演集 45th 46 2011年03月18日 [査読無し][通常論文]
  • Takeshi Yamada, Kae Kikuchi, Toshihiro Yamauchi, Koji Shiraishi, Tsukasa Ito, Satoshi Okabe, Akira Hiraishi, Akiyoshi Ohashi, Hideki Harada, Yoichi Kamagata, Kazunori Nakamura, Yuji Sekiguchi APPLIED AND ENVIRONMENTAL MICROBIOLOGY 77 (6) 2081 -2087 2011年03月 [査読無し][通常論文]
     
    A filamentous bulking of a methanogenic granular sludge caused by uncultured filamentous bacteria of the candidate phylum KSB3 in an upflow anaerobic sludge blanket (UASB) system has been reported. To characterize the physiological traits of the filaments, a polyphasic approach consisting of rRNA-based activity monitoring of the KSB3 filaments using the RNase H method and substrate uptake profiling using microautoradiography combined with fluorescence in situ hybridization (MAR-FISH) was conducted. On the basis of rRNA-based activity, the monitoring of a full-scale UASB reactor operated continuously revealed that KSB3 cells became active and predominant (up to 54% of the total 16S rRNA) in the sludge when the carbohydrate loading to the system increased. Batch experiments with a short incubation of the sludge with maltose, glucose, fructose, and maltotriose at relatively low concentrations (approximately 0.1 mM) in the presence of yeast extract also showed an increase in KSB3 rRNA levels under anaerobic conditions. MAR-FISH confirmed that the KSB3 cells took up radioisotopic carbons from [(14)C] maltose and [(14)C] glucose under the same incubation conditions in the batch experiments. These results suggest that one of the important ecophysiological characteristics of KSB3 cells in the sludge is carbohydrate degradation in wastewater and that high carbohydrate loadings may trigger an outbreak of KSB3 bacteria, causing sludge bulking in UASB systems.
  • Takeshi Yamada, Kae Kikuchi, Toshihiro Yamauchi, Koji Shiraishi, Tsukasa Ito, Satoshi Okabe, Akira Hiraishi, Akiyoshi Ohashi, Hideki Harada, Yoichi Kamagata, Kazunori Nakamura, Yuji Sekiguchi APPLIED AND ENVIRONMENTAL MICROBIOLOGY 77 (6) 2081 -2087 2011年03月 [査読無し][通常論文]
     
    A filamentous bulking of a methanogenic granular sludge caused by uncultured filamentous bacteria of the candidate phylum KSB3 in an upflow anaerobic sludge blanket (UASB) system has been reported. To characterize the physiological traits of the filaments, a polyphasic approach consisting of rRNA-based activity monitoring of the KSB3 filaments using the RNase H method and substrate uptake profiling using microautoradiography combined with fluorescence in situ hybridization (MAR-FISH) was conducted. On the basis of rRNA-based activity, the monitoring of a full-scale UASB reactor operated continuously revealed that KSB3 cells became active and predominant (up to 54% of the total 16S rRNA) in the sludge when the carbohydrate loading to the system increased. Batch experiments with a short incubation of the sludge with maltose, glucose, fructose, and maltotriose at relatively low concentrations (approximately 0.1 mM) in the presence of yeast extract also showed an increase in KSB3 rRNA levels under anaerobic conditions. MAR-FISH confirmed that the KSB3 cells took up radioisotopic carbons from [(14)C] maltose and [(14)C] glucose under the same incubation conditions in the batch experiments. These results suggest that one of the important ecophysiological characteristics of KSB3 cells in the sludge is carbohydrate degradation in wastewater and that high carbohydrate loadings may trigger an outbreak of KSB3 bacteria, causing sludge bulking in UASB systems.
  • 佐藤久, 池田真之, 高橋正宏, 岡部聡, 笹川学, 中原禎仁 環境浄化技術 10 (1) 85 -90 2011年02月01日 [査読無し][通常論文]
  • Daisuke Sano, Unai Perez-Sautu, Susana Guix, Rosa Maria Pinto, Takayuki Miura, Satoshi Okabe, Albert Bosch APPLIED AND ENVIRONMENTAL MICROBIOLOGY 77 (3) 1111 -1114 2011年02月 [査読無し][通常論文]
     
    Human sapoviruses (SaVs) were quantified and characterized in an 18-month survey conducted along the Llobregat river catchment area in Spain. Sample types included freshwater, untreated and treated wastewater, and drinking water. All genogroups were recovered, and a seasonal distribution was observed. This is the first report of SaV quantification and genotyping in the environment outside Japan.
  • Akira Hafuka, Kenji Sakaida, Hisashi Satoh, Masahiro Takahashi, Yoshimasa Watanabe, Satoshi Okabe BIORESOURCE TECHNOLOGY 102 (3) 3551 -3553 2011年02月 [査読無し][通常論文]
     
    We investigated the effects of different feeding regimens (1-pulse, stepwise, and continuous) of fermented food-waste liquid on polyhydroxybutyrate (PHB) production. The fermentation liquid was filtered with a membrane filter (pore size, 0.45 mu m) to remove anaerobic microorganisms and solids and used as a carbon source for Cupriavidus necator. One-pulse feeding yielded the highest cell concentration of C. necator. However, the PHB concentration was higher in the stepwise- and continuous-feeding regimens. Therefore, the continuous-feeding regimen was used for continuous PHB production. PHB could be produced over 259 h (8 draw-fill cycles) with a maximal PHB content of 87%, but the PHB concentration and content decreased with an increase in the operation time. (C) 2010 Elsevier Ltd. All rights reserved.
  • Daisuke Sano, Unai Perez-Sautu, Susana Guix, Rosa Maria Pinto, Takayuki Miura, Satoshi Okabe, Albert Bosch APPLIED AND ENVIRONMENTAL MICROBIOLOGY 77 (3) 1111 -1114 2011年02月 [査読無し][通常論文]
     
    Human sapoviruses (SaVs) were quantified and characterized in an 18-month survey conducted along the Llobregat river catchment area in Spain. Sample types included freshwater, untreated and treated wastewater, and drinking water. All genogroups were recovered, and a seasonal distribution was observed. This is the first report of SaV quantification and genotyping in the environment outside Japan.
  • Akira Hafuka, Kenji Sakaida, Hisashi Satoh, Masahiro Takahashi, Yoshimasa Watanabe, Satoshi Okabe BIORESOURCE TECHNOLOGY 102 (3) 3551 -3553 2011年02月 [査読無し][通常論文]
     
    We investigated the effects of different feeding regimens (1-pulse, stepwise, and continuous) of fermented food-waste liquid on polyhydroxybutyrate (PHB) production. The fermentation liquid was filtered with a membrane filter (pore size, 0.45 mu m) to remove anaerobic microorganisms and solids and used as a carbon source for Cupriavidus necator. One-pulse feeding yielded the highest cell concentration of C. necator. However, the PHB concentration was higher in the stepwise- and continuous-feeding regimens. Therefore, the continuous-feeding regimen was used for continuous PHB production. PHB could be produced over 259 h (8 draw-fill cycles) with a maximal PHB content of 87%, but the PHB concentration and content decreased with an increase in the operation time. (C) 2010 Elsevier Ltd. All rights reserved.
  • Sunja Cho, Naoki Fujii, Taeho Lee, Satoshi Okabe BIORESOURCE TECHNOLOGY 102 (2) 652 -659 2011年01月 [査読無し][通常論文]
     
    Up-flow oxygen-controlled biofilm reactors equipped with a non-woven fabric support were used as a single reactor system for autotrophic nitrogen removal based on a combined partial nitrification and anaerobic ammonium oxidation (anammox) reaction. The up-flow biofilm reactors were initiated as either a partial nitrifying reactor or an anammox reactor, respectively, and simultaneous partial nitrification and anammox was established by careful control of the aeration rate. The combined partial nitrification and anammox reaction was successfully developed in both biofilm reactors without additional biomass inoculation. The reactor initiated as the anammox reactor gave a slightly higher and more stable mean nitrogen removal rate of 0.35 (+/- 0.19) kg-N m(-3) d(-1) than the reactor initiated as the partial nitrifying reactor (0.23 (+/- 0.16) kg-N m(-3) d(-1)). FISH analysis revealed that the biofilm in the reactor started as the anammox reactor were composed of anammox bacteria located in inner anoxic layers that were surrounded by surface aerobic AOB layers, whereas AOB and anammox bacteria were mixed without a distinguishable niche in the biofilm in the reactor started as the partial nitrifying reactor. However, it was difficult to efficiently maintain the stable partial nitrification owing to inefficient aeration in the reactor, which is a key to development of the combined partial nitrification and anammox reaction in a single biofilm reactor. (C) 2010 Elsevier Ltd. All rights reserved.
  • Kyungmi Chung, Itto Fujiki, Satoshi Okabe BIORESOURCE TECHNOLOGY 102 (1) 355 -360 2011年01月 [査読無し][通常論文]
     
    A two-chamber MFC system was operated continuously for more than 500 days to evaluate effects of biofilm and chemical scale formation on the cathode electrode on power generation. A stable power density of 0.57 W/m(2) was attained after 200 days operation. However, the power density decreased drastically to 0.2 W/m(2) after the cathodic biofilm and chemical scale were removed. As the cathodic biofilm and chemical scale partially accumulated on the cathode, the power density gradually recovered with time. Microbial community structure of the cathodic biofilm was analyzed based on 16S rRNA clone libraries. The clones closely related to Xanthomonadaceae bacterium and Xanthomonas sp. in the Gammaproteobacteria subdivision were most frequently retrieved from the cathodic biofilm. Results of the SEM-EDX analysis revealed that the cation species (Na(+) and Ca(2+)) were main constituents of chemical scale, indicating that these cations diffused from the anode chamber through the Nation membrane. However, an excess accumulation of the biofilm and chemical scale on the cathode exhibited adverse effects on the power generation due to a decrease in the active cathode surface area and an increase in diffusion resistance for oxygen. Thus, it is important to properly control the formation of chemical scale and biofilm on the cathode during long-term operation. (C) 2010 Elsevier Ltd. All rights reserved.
  • Satoshi Okabe, Hisashi Satoh, Tomonori Kindaichi METHODS IN ENZYMOLOGY, VOL 46: RESEARCH ON NITRIFICATION AND RELATED PROCESSES, PT B 496 163 -184 2011年 [査読無し][通常論文]
     
    This chapter aims to highlight the great potential of the combined use of microautoradiography (MAR) combined with fluorescent in situ hybridization (FISH) and microsensor technology in studies of complex multispecies nitrifying biofilms. The combination of FISH and microsensor technology is a powerful and reliable tool to link the spatial organization of microbial communities and their in situ function at community levels. MAR-FISH can be used to simultaneously examine the 16S rRNA-based phylogenetic identity and specific metabolic activity of cultivable or uncultivable microorganisms within complex microbial communities at a single-cell level. Information obtained at both resolution levels must be combined to draw a clear picture of a complex multispecies biofilm ecosystem. In addition, ecophysiological interactions among community members in complex multispecies biofilms can be investigated by tracing the fate of radiolabeled [(14)C] atom incorporated in nitrifying bacteria with MAR-FISH. The structure, function, and ecophysiological interactions among community members in complex multispecies nitrifying biofilms will be illustrated as an example of the combined use of MAR-FISH and microsensor technology.
  • Analyses of three dominant membrane proteins from anammox planctomycete Candidatus 'Brocadia sinica'
    Journal of Environmental Biotechnology 11 (1-2) 1 2011年 [査読無し][通常論文]
  • Sunja Cho, Naoki Fujii, Taeho Lee, Satoshi Okabe BIORESOURCE TECHNOLOGY 102 (2) 652 -659 2011年01月 [査読無し][通常論文]
     
    Up-flow oxygen-controlled biofilm reactors equipped with a non-woven fabric support were used as a single reactor system for autotrophic nitrogen removal based on a combined partial nitrification and anaerobic ammonium oxidation (anammox) reaction. The up-flow biofilm reactors were initiated as either a partial nitrifying reactor or an anammox reactor, respectively, and simultaneous partial nitrification and anammox was established by careful control of the aeration rate. The combined partial nitrification and anammox reaction was successfully developed in both biofilm reactors without additional biomass inoculation. The reactor initiated as the anammox reactor gave a slightly higher and more stable mean nitrogen removal rate of 0.35 (+/- 0.19) kg-N m(-3) d(-1) than the reactor initiated as the partial nitrifying reactor (0.23 (+/- 0.16) kg-N m(-3) d(-1)). FISH analysis revealed that the biofilm in the reactor started as the anammox reactor were composed of anammox bacteria located in inner anoxic layers that were surrounded by surface aerobic AOB layers, whereas AOB and anammox bacteria were mixed without a distinguishable niche in the biofilm in the reactor started as the partial nitrifying reactor. However, it was difficult to efficiently maintain the stable partial nitrification owing to inefficient aeration in the reactor, which is a key to development of the combined partial nitrification and anammox reaction in a single biofilm reactor. (C) 2010 Elsevier Ltd. All rights reserved.
  • Kyungmi Chung, Itto Fujiki, Satoshi Okabe BIORESOURCE TECHNOLOGY 102 (1) 355 -360 2011年01月 [査読無し][通常論文]
     
    A two-chamber MFC system was operated continuously for more than 500 days to evaluate effects of biofilm and chemical scale formation on the cathode electrode on power generation. A stable power density of 0.57 W/m(2) was attained after 200 days operation. However, the power density decreased drastically to 0.2 W/m(2) after the cathodic biofilm and chemical scale were removed. As the cathodic biofilm and chemical scale partially accumulated on the cathode, the power density gradually recovered with time. Microbial community structure of the cathodic biofilm was analyzed based on 16S rRNA clone libraries. The clones closely related to Xanthomonadaceae bacterium and Xanthomonas sp. in the Gammaproteobacteria subdivision were most frequently retrieved from the cathodic biofilm. Results of the SEM-EDX analysis revealed that the cation species (Na(+) and Ca(2+)) were main constituents of chemical scale, indicating that these cations diffused from the anode chamber through the Nation membrane. However, an excess accumulation of the biofilm and chemical scale on the cathode exhibited adverse effects on the power generation due to a decrease in the active cathode surface area and an increase in diffusion resistance for oxygen. Thus, it is important to properly control the formation of chemical scale and biofilm on the cathode during long-term operation. (C) 2010 Elsevier Ltd. All rights reserved.
  • Satoshi Okabe, Hisashi Satoh, Tomonori Kindaichi Methods in Enzymology 496 163 -184 2011年 [査読無し][通常論文]
     
    This chapter aims to highlight the great potential of the combined use of microautoradiography (MAR) combined with fluorescent in situ hybridization (FISH) and microsensor technology in studies of complex multispecies nitrifying biofilms. The combination of FISH and microsensor technology is a powerful and reliable tool to link the spatial organization of microbial communities and their in situ function at community levels. MAR-FISH can be used to simultaneously examine the 16S rRNA-based phylogenetic identity and specific metabolic activity of cultivable or uncultivable microorganisms within complex microbial communities at a single-cell level. Information obtained at both resolution levels must be combined to draw a clear picture of a complex multispecies biofilm ecosystem. In addition, ecophysiological interactions among community members in complex multispecies biofilms can be investigated by tracing the fate of radiolabeled [14C] atom incorporated in nitrifying bacteria with MAR-FISH. The structure, function, and ecophysiological interactions among community members in complex multispecies nitrifying biofilms will be illustrated as an example of the combined use of MAR-FISH and microsensor technology. © 2011 Elsevier Inc. All rights reserved.
  • Analyses of three dominant membrane proteins from anammox planctomycete Candidatus 'Brocadia sinica'
    Journal of Environmental Biotechnology 11 (1-2) 1 2011年 [査読無し][通常論文]
  • Satoshi Okabe, Mamoru Oshiki, Yoichi Kamagata, Nobuyasu Yamaguchi, Masanori Toyofuku, Yutaka Yawata, Yosuke Tashiro, Nobuhiko Nomura, Hiroyuki Ohta, Moriya Ohkuma, Akira Hiraishi, Kiwamu Minamisawa MICROBES AND ENVIRONMENTS 25 (4) 230 -240 2010年12月 [査読無し][通常論文]
     
    Ribosomal RNA (rRNA) sequence-based molecular techniques emerged in the late 1980s, which completely changed our general view of microbial life. Coincidentally, the Japanese Society of Microbial Ecology (JSME) was founded, and its official journal "Microbes and Environments (M&E)" was launched, in 1985. Thus, the past 25 years have been an exciting and fruitful period for M&E readers and microbiologists as demonstrated by the numerous excellent papers published in M&E. In this minireview, recent progress made in microbial ecology and related fields is summarized, with a special emphasis on 8 landmark areas; the cultivation of uncultured microbes, in situ methods for the assessment of microorganisms and their activities, biofilms, plant microbiology, chemolithotrophic bacteria in early volcanic environments, symbionts of animals and their ecology, wastewater treatment microbiology, and the biodegradation of hazardous organic compounds.
  • 新家子 香織, 押木 守, 佐藤 久, 岡部 聡 日本微生物生態学会講演要旨集 (26) 2010年11月23日 [査読無し][通常論文]
  • 羽深昭, 谷山拓生, 山田幸司, 高橋正宏, 岡部聡, 佐藤久 環境工学研究フォーラム講演集 47th 184 -186 2010年11月12日 [査読無し][通常論文]
  • 宮崎悠爾, 押木守, 佐藤久, 高橋正宏, 岡部聡 環境工学研究フォーラム講演集 47th 4 -6 2010年11月12日 [査読無し][通常論文]
  • Thithiwat May, Akinobu Ito, Satoshi Okabe MOLECULAR GENETICS AND GENOMICS 284 (5) 333 -342 2010年11月 [査読無し][通常論文]
     
    The ecological role of horizontal gene transfer within biofilms has been recently investigated, and it has been reported that conjugation directly induces bacteria to form biofilms via expression of conjugative pili. In this report, we described the contribution of bacterial conjugation during biofilm formation by Escherichia coli harboring a natural IncF conjugative F plasmid (F+). We showed that cell-to-cell pili interactions through the homosexual mating-pair formation among F+ x F+ cells (namely, F- phenocopy phenomenon) promote E. coli biofilm formation at the early development stage. The presence of F+ x F+ population is the result from heterogeneity within biofilms leading to sessile bacteria that grow at different rates, in which the late-stationary phase cells acted as F- phenocopy cells. According to global transcriptional analysis, the biofilm lifestyle shared similar gene expression pattern with F- phenocopies. F- phenocopy cells expressed specific sets of chromosomal genes (e.g., genes for general stress response and two-component systems) that control the regulation regions of F transfer operon by blocking surface exclusion proteins and DNA transfer machineries. However, mating-pair proteins were stabilized and consequently promoted F+ x F+ pili assembly. Thus, F- phenocopy phenomenon is an effective adaptive behavior of bacterial cells during biofilm formation.
  • Thithiwat May, Akinobu Ito, Satoshi Okabe MOLECULAR GENETICS AND GENOMICS 284 (5) 333 -342 2010年11月 [査読無し][通常論文]
     
    The ecological role of horizontal gene transfer within biofilms has been recently investigated, and it has been reported that conjugation directly induces bacteria to form biofilms via expression of conjugative pili. In this report, we described the contribution of bacterial conjugation during biofilm formation by Escherichia coli harboring a natural IncF conjugative F plasmid (F+). We showed that cell-to-cell pili interactions through the homosexual mating-pair formation among F+ x F+ cells (namely, F- phenocopy phenomenon) promote E. coli biofilm formation at the early development stage. The presence of F+ x F+ population is the result from heterogeneity within biofilms leading to sessile bacteria that grow at different rates, in which the late-stationary phase cells acted as F- phenocopy cells. According to global transcriptional analysis, the biofilm lifestyle shared similar gene expression pattern with F- phenocopies. F- phenocopy cells expressed specific sets of chromosomal genes (e.g., genes for general stress response and two-component systems) that control the regulation regions of F transfer operon by blocking surface exclusion proteins and DNA transfer machineries. However, mating-pair proteins were stabilized and consequently promoted F+ x F+ pili assembly. Thus, F- phenocopy phenomenon is an effective adaptive behavior of bacterial cells during biofilm formation.
  • 佐藤久, 日向寺崇文, 高橋正宏, 岡部聡 土木学会年次学術講演会講演概要集(CD-ROM) 65th ROMBUNNO.VII-176 2010年08月05日 [査読無し][通常論文]
  • 木村善一郎, 岡部聡 日本水環境学会年会講演集 44th 507 2010年03月15日 [査読無し][通常論文]
  • 伊藤皓亮, 木村善一郎, CHUNG Kyungmi, 岡部聡 日本水環境学会年会講演集 44th 512 2010年03月15日 [査読無し][通常論文]
  • 佐藤久, 高橋慶考, 山田陽平, 高橋正宏, 岡部聡 日本水環境学会年会講演集 44th 396 2010年03月15日 [査読無し][通常論文]
  • Sunja Cho, Yoshitaka Takahashi, Naoki Fujii, Yohei Yamada, Hisashi Satoh, Satoshi Okabe CHEMOSPHERE 78 (9) 1129 -1135 2010年02月 [査読無し][通常論文]
     
    We investigated nitrogen removal performance and responsible microbial community in an anaerobic up-flow granular bed anammox reactor. The anammox reactor was operated more than 1 year. Biomass in the reactor formed granules after about 2 months of operation, and a sufficient amount of the granules was retained in the reactor with a metallic net to avoid biomass washout during the entire operation. The average diameter of the granules was 3.6 mm at day 310. After 8 months of operation, stable nitrogen removal (60%) was achieved at an average total inorganic nitrogen removal rate of 14 kg-N m(-3) d(-1). The phylogenetic analysis and fluorescence in situ hybridization results revealed that the anammox granules consisted of mono species of anammox bacteria, "Candidatus Brocadia-like species", affiliated with "Candidatus Brocadia anammoxidans" with 16S rRNA gene sequence similarity of 95.7%. The relative abundance of the anammox bacteria in the granules was more than 80% of the total bacteria stained with 4',6-diamidino-2-phenylindole. The anammox bacteria were present throughout the granules whereas the other bacterial groups. Chloroflexi-like filamentous bacteria and betaproteobacterial ammonia-oxidizing bacteria, were mainly present on the surface of the anammox granules and around the anammox bacterial clusters. The in situ anammox activity was detected mainly from near the surface of granules to the upper 800 pm of the granules with microsensors. The granular anammox biomass tolerated higher concentrations of nitrite (400 mg-N L(-1)) than did the homogenized biomass (200 mg-N L(-1)) probably due to substrate diffusion limitation. (C) 2009 Elsevier Ltd. All rights reserved.
  • Sunja Cho, Yoshitaka Takahashi, Naoki Fujii, Yohei Yamada, Hisashi Satoh, Satoshi Okabe CHEMOSPHERE 78 (9) 1129 -1135 2010年02月 [査読無し][通常論文]
     
    We investigated nitrogen removal performance and responsible microbial community in an anaerobic up-flow granular bed anammox reactor. The anammox reactor was operated more than 1 year. Biomass in the reactor formed granules after about 2 months of operation, and a sufficient amount of the granules was retained in the reactor with a metallic net to avoid biomass washout during the entire operation. The average diameter of the granules was 3.6 mm at day 310. After 8 months of operation, stable nitrogen removal (60%) was achieved at an average total inorganic nitrogen removal rate of 14 kg-N m(-3) d(-1). The phylogenetic analysis and fluorescence in situ hybridization results revealed that the anammox granules consisted of mono species of anammox bacteria, "Candidatus Brocadia-like species", affiliated with "Candidatus Brocadia anammoxidans" with 16S rRNA gene sequence similarity of 95.7%. The relative abundance of the anammox bacteria in the granules was more than 80% of the total bacteria stained with 4',6-diamidino-2-phenylindole. The anammox bacteria were present throughout the granules whereas the other bacterial groups. Chloroflexi-like filamentous bacteria and betaproteobacterial ammonia-oxidizing bacteria, were mainly present on the surface of the anammox granules and around the anammox bacterial clusters. The in situ anammox activity was detected mainly from near the surface of granules to the upper 800 pm of the granules with microsensors. The granular anammox biomass tolerated higher concentrations of nitrite (400 mg-N L(-1)) than did the homogenized biomass (200 mg-N L(-1)) probably due to substrate diffusion limitation. (C) 2009 Elsevier Ltd. All rights reserved.
  • Satoshi Okabe, Mamoru Oshiki, Yoichi Kamagata, Nobuyasu Yamaguchi, Masanori Toyofuku, Yutaka Yawata, Yosuke Tashiro, Nobuhiko Nomura, Hiroyuki Ohta, Moriya Ohkuma, Akira Hiraishi, Kiwamu Minamisawa Microbes and environments 25 (4) 230 -40 2010年 [査読有り][通常論文]
     
    Ribosomal RNA (rRNA) sequence-based molecular techniques emerged in the late 1980s, which completely changed our general view of microbial life. Coincidentally, the Japanese Society of Microbial Ecology (JSME) was founded, and its official journal "Microbes and Environments (M&E)" was launched, in 1985. Thus, the past 25 years have been an exciting and fruitful period for M&E readers and microbiologists as demonstrated by the numerous excellent papers published in M&E. In this minireview, recent progress made in microbial ecology and related fields is summarized, with a special emphasis on 8 landmark areas; the cultivation of uncultured microbes, in situ methods for the assessment of microorganisms and their activities, biofilms, plant microbiology, chemolithotrophic bacteria in early volcanic environments, symbionts of animals and their ecology, wastewater treatment microbiology, and the biodegradation of hazardous organic compounds.
  • Kyungmi Chung, Satoshi Okabe Biotechnology and bioengineering 104 (5) 901 -10 2009年12月01日 [査読有り][通常論文]
     
    The microbial communities associated with electrodes in closed and open circuit microbial fuel cells (MFCs) fed with glucose were analyzed by 16S rRNA approach and compared. The comparison revealed that bacteria affiliated with the Aeromonas sp. within the Gammaproteobacteria constituted the major population in the closed circuit MFC (harvesting electricity) and considered to play important roles in current generation. We, therefore, attempted to isolate the dominant bacteria from the anode biofilm, successfully isolated a Fe (III)-reducing bacterium phylogenetically related to Aeromonas sp. and designated as strain ISO2-3. The isolated strain ISO2-3 could grow and concomitantly produce current (max. 0.24 A/m(2)) via oxidation of glucose or hydrogen with an electrode serving as the sole electron acceptor. The strain could ferment glucose, but generate less electrical current. Cyclic voltammetry supported the strain ISO2-3 was electrically active and likely to transfer electrons to the electrode though membrane-associated compounds (most likely c-type cytochrome). This mechanism requires intimate contact with the anode surface. Scanning electron microscopy revealed that the strain ISO2-3 developed multiplayer biofilms on the anode surface and also produced anchor-like filamentous appendages (most likely pili) that may promote long-range electron transport across the thick biofilm.
  • 池田真之, 佐藤久, 高橋正宏, 岡部聡, 中原禎仁, 笹川学 環境工学研究フォーラム講演集 46th 103 -104 2009年11月27日 [査読無し][通常論文]
  • 佐藤久, 池田真之, 高橋正宏, 岡部聡, 中原禎仁, 笹川学 用水と廃水 51 (11) 925 -932 2009年11月01日 [査読無し][通常論文]
  • Thithiwat May, Akinobu Ito, Satoshi Okabe Antimicrobial agents and chemotherapy 53 (11) 4628 -39 2009年11月 [査読有り][通常論文]
     
    Biofilms gain resistance to various antimicrobial agents, and the presence of antibiotic resistance genes is thought to contribute to a biofilm-mediated antibiotic resistance. Here we showed the interplay between the tetracycline resistance efflux pump TetA(C) and the ampicillin resistance gene (bla(TEM-1)) in biofilms of Escherichia coli harboring pBR322 in the presence of the mixture of ampicillin and tetracycline. E. coli in the biofilms could obtain the high-level resistance to ampicillin, tetracycline, penicillin, erythromycin, and chloramphenicol during biofilm development and maturation as a result of the interplay between the marker genes on the plasmids, the increase of plasmid copy number, and consequently the induction of the efflux systems on the bacterial chromosome, especially the EmrY/K and EvgA/S pumps. In addition, we characterized the overexpression of the TetA(C) pump that contributed to osmotic stress response and was involved in the induction of capsular colanic acid production, promoting formation of mature biofilms. However, this investigated phenomenon was highly dependent on the addition of the subinhibitory concentrations of antibiotic mixture, and the biofilm resistance behavior was limited to aminoglycoside antibiotics. Thus, marker genes on plasmids played an important role in both resistance of biofilm cells to antibiotics and in formation of mature biofilms, as they could trigger specific chromosomal resistance mechanisms to confer a high-level resistance during biofilm formation.
  • Hisashi Satoh, Mitsunori Odagiri, Tsukasa Ito, Satoshi Okabe WATER RESEARCH 43 (18) 4729 -4739 2009年10月 [査読無し][通常論文]
     
    Microbially induced concrete corrosion (MICC) caused by sulfuric acid attack in sewer systems has been a serious problem for a long time. A better understanding of microbial community structures of sulfate-reducing bacteria (SRB) and sulfur-oxidizing bacteria (SOB) and their in situ activities is essential for the efficient control of MICC. in this study, the microbial community structures and the in situ hydrogen sulfide production and consumption rates within biofilms and corroded materials developed on mortar specimens placed in a corroded manhole was investigated by culture-independent 16S rRNA gene-based molecular techniques and microsensors for hydrogen sulfide, oxygen, pH and the oxidation-reduction potential. The dark-gray gel-like biofilm was developed in the bottom (from the bottom to 4 cm) and the middle (4-20 cm from the bottom of the manhole) parts of the mortar specimens. White filamentous biofilms covered the gel-like biofilm in the middle part. The mortar specimens placed in the upper part (30 cm above the bottom of the manhole) were corroded. The 16S rRNA gene-cloning analysis revealed that one clone retrieved from the bottom biofilm sample was related to an SRB, 12 clones and 6 clones retrieved from the middle biofilm and the corroded material samples, respectively, were related to SOB. In situ hybridization results showed that the SRB were detected throughout the bottom biofilm and filamentous SOB cells were mainly detected in the upper oxic layer of the middle biofilm. Microsensor measurements demonstrated that hydrogen sulfide was produced in and diffused out of the bottom biofilms. In contrast, in the middle biofilm the hydrogen sulfide produced in the deeper parts of the biofilm was oxidized in the upper filamentous biofilm. pH was around 3 in the corroded materials developed in the upper part of the mortar specimens. Therefore, it can be concluded that hydrogen sulfide provided from the bottom biofilms and the sludge settling tank was emitted to the sewer atmosphere, then oxidized to corrosive compounds in the upper and middle parts of the manhole, and only the upper part of the mortar specimens were corroded, because in the middle part of the manhole the generated corrosive compounds (e.g., sulfuric acid) was reduced in the deeper parts of the biofilm. (C) 2009 Elsevier Ltd. All rights reserved.
  • 羽深昭, 佐藤久, 高橋正宏, 岡部聡 日本水環境学会シンポジウム講演集 12th 168 -169 2009年09月14日 [査読無し][通常論文]
  • Koji Kawata, Masato Osawa, Satoshi Okabe ENVIRONMENTAL SCIENCE & TECHNOLOGY 43 (15) 6046 -6051 2009年08月 [査読無し][通常論文]
     
    Although it has been reported that silver nanoparticles (Ag-NPs) have strong acute toxic effects to various cultured the toxic effects at noncytotoxic doses are still unknown. We, therefore, evaluated in vitro toxicity of doses in human hepatoma cell line, HepG2, based on cell viability assay, micronucleus test, and DNA microarray We also used polystyrene nanoparticles (PS-NPs) and silver carbonate (Ag(2)CO(3)) as test materials to compare the toxic with respect to different raw chemical composition and form of silver. The cell viability assay demonstrated that Ag-NPs accelerated cell proliferation at low doses (<0.5 mg/L), was supported by the DNA microarray analysis showing significant induction of genes associated with cell cycle progression. However, only Ag-NPs exposure exhibited a significant cytotoxicity at higher doses (> 1.0 mg/L) and abnormal cellular morphology, displaying cellular shrinkage and acquisition of an irregular shape. In addition, only exposure increased the frequency of micronucleus formation up to 47.9 +/- 3.2% of binucleated cells, suggesting that appear to cause much stronger damages to chromosome than PS-NPs and ionic Ag(+). Cysteine, a strong ionic Ag(+) only partially abolished the formation of micronuclei mediated by Ag-NPs and changed the gene expression, indicating that ionic Ag(+) derived from Ag-NPs could not fully explain these biological actions, Based on these discussions, it is concluded that both "nanosized particle of Ag" as well as "ionic Ag(+)" contribute to the toxic effects of Ag-NPs.
  • Akinobu Ito, Thithiwat May, Asami Taninchi, Koji Kawata, Satoshi Okabe BIOTECHNOLOGY AND BIOENGINEERING 103 (5) 975 -983 2009年08月 [査読無し][通常論文]
     
    Although importance of the rpoS gene on biofilm formation by Escherichia coli has been suggested, there his not been any report showing where the rpoS is expressed during biofilm formation process. Since physiological state of the cells in the biofilms is considerably heterogeneous, the expression of the rpoS gene must be heterogeneous. In this study, in situ spatial expression of the rpoS gene during biofilm formation was investigated with an rpoS-gfp transcriptional fusion mutant strain. A ribosomal binding site and a gene encoding a green fluorescent protein were introduced into the downstream of the rpoS gene, which enabled us to observe the in situ spatial expression of the rpoS gene during biofilm formation processes without any disturbance of the rpoS expression. In the early stages of the biofilm formation process, the rpoS gene was expressed in the most of the cells. On the other hand, the rpoS expression was observed only at the outside of the biofilms during the late stages of the biofilm formation process. The in situ spatial expression of the rpoS gene in the biofilm was verified by quantifying the expression levels of the rpoS at the outside and the inside of the biofilms with the real time RTPCR. In addition, global gene expression analysis was performed with DNA microarray to investigate physiological difference between the outside and the inside of the biofilms. This heterogeneous rpoS expression profile suggested that the cells at the outside of the biofilm need to express the rpoS to shift the physiological state to the stationary growth mode such as induction of various stress responses and suppression of the motility. Biotechnol. Bioeng. 2009;103: 975-983. (C) 2009 Wiley Periodicals, Inc.
  • Feng Jiang, Derek Hoi-wai Leung, Shiyu Li, Guang-Hao Chen, Satoshi Okabe, Mark C. M. van Loosdrecht WATER RESEARCH 43 (13) 3187 -3198 2009年07月 [査読無し][通常論文]
     
    This study developed a new sewer biofilm model to simulate the pollutant transformation and biofilm variation in sewers under aerobic, anoxic and anaerobic conditions. The biofilm model can describe the activities of heterotrophic, autotrophic, and sulfate-reducing bacteria (SRB) in the biofilm as well as the variations in biofilm thickness, the spatial profiles of SRB population and biofilm density. The model can describe dynamic biofilm growth, multiple biomass evolution and competitions among organic oxidation, denitrification, nitrification, sulfate reduction and sulfide oxidation in a heterogeneous biofilm growing in a sewer. The model has been extensively verified by three different approaches, including direct verification by measurement of the spatial concentration profiles of dissolved oxygen, nitrate, ammonia, and hydrogen sulfide in sewer biofilm. The spatial distribution profile of SRB in sewer biofilm was determined from the fluorescent in situ hybridization (FISH) images taken by a confocal laser scanning microscope (CLSM) and were predicted well by the model. (C) 2009 Elsevier Ltd. All rights reserved.
  • Kyungmi Chung, Satoshi Okabe APPLIED MICROBIOLOGY AND BIOTECHNOLOGY 83 (5) 965 -977 2009年07月 [査読無し][通常論文]
     
    A mediator-less three-stage two-chamber microbial fuel cell (MFC) system was developed and operated continuously for more than 1.5 years to evaluate continuous power generation while treating artificial wastewater containing glucose (10 mM) concurrently. A stable power density of 28 W/m(3) was attained with an anode hydraulic retention time of 4.5 h and phosphate buffer as the cathode electrolyte. An overall dissolved organic carbon removal ratio was about 85%, and coulombic efficiency was about 46% in this MFC system. We also analyzed the microbial community structure of anode biofilms in each MFC. Since the environment in each MFC was different due to passing on the products to the next MFC in series, the microbial community structure was different accordingly. The anode biofilm in the first MFC consisted mainly of bacteria belonging to the Gammaproteobacteria, identified as Aeromonas sp., while the Firmicutes dominated the anode biofilms in the second and third MFCs that were mainly fed with acetate. Cyclic voltammetric results supported the presence of a redox compound(s) associated with the anode biofilm matrix, rather than mobile (dissolved) forms, which could be responsible for the electron transfer to the anode. Scanning electron microscopy revealed that the anode biofilms were comprised of morphologically different cells that were firmly attached on the anode surface and interconnected each other with anchor-like filamentous appendages, which might support the results of cyclic voltammetry.
  • Akinobu Ito, Asami Taniuchi, Thithiwat May, Koji Kawata, Satoshi Okabe APPLIED AND ENVIRONMENTAL MICROBIOLOGY 75 (12) 4093 -4100 2009年06月 [査読無し][通常論文]
     
    Biofilms are considered to be highly resistant to antimicrobial agents. Several mechanisms have been proposed to explain this high resistance of biofilms, including restricted penetration of antimicrobial agents into biofilms, slow growth owing to nutrient limitation, expression of genes involved in the general stress response, and emergence of a biofilm-specific phenotype. However, since combinations of these factors are involved in most biofilm studies, it is still difficult to fully understand the mechanisms of biofilm resistance to antibiotics. In this study, the antibiotic susceptibility of Escherichia coli cells in biofilms was investigated with exclusion of the effects of the restricted penetration of antimicrobial agents into biofilms and the slow growth owing to nutrient limitation. Three different antibiotics, ampicillin (100 mu g/ml), kanamycin (25 mu g/ml), and ofloxacin (10 mu g/ml), were applied directly to cells in the deeper layers of mature biofilms that developed in flow cells after removal of the surface layers of the biofilms. The results of the antibiotic treatment analyses revealed that ofloxacin and kanamycin were effective against biofilm cells, whereas ampicillin did not kill the cells, resulting in regrowth of the biofilm after the ampicillin treatment was discontinued. LIVE/DEAD staining revealed that a small fraction of resistant cells emerged in the deeper layers of the mature biofilms and that these cells were still alive even after 24 h of ampicillin treatment. Furthermore, to determine which genes in the biofilm cells are induced, allowing increased resistance to ampicillin, global gene expression was analyzed at different stages of biofilm formation, the attachment, colony formation, and maturation stages. The results showed that significant changes in gene expression occurred during biofilm formation, which were partly induced by rpoS expression. Based on the experimental data, it is likely that the observed resistance of biofilms can be attributed to formation of ampicillin-resistant subpopulations in the deeper layers of mature biofilms but not in young colony biofilms and that the production and resistance of the subpopulations were aided by biofilm-specific phenotypes, like slow growth and induction of rpoS-mediated stress responses.
  • 佐藤久, 池田真之, 高橋正宏, 岡部聡, 中原禎仁, 笹川学 日本水環境学会年会講演集 43rd 253 2009年03月16日 [査読無し][通常論文]
  • 羽深昭, 佐藤久, 高橋正宏, 岡部聡 日本水環境学会年会講演集 43rd 623 2009年03月16日 [査読無し][通常論文]
  • 木村善一郎, 藤木一到, 岡部聡 日本微生物生態学会講演要旨集 25th YOSHI74 2009年 [査読無し][通常論文]
  • 伊藤皓亮, 木村善一郎, 鄭景美, 岡部聡 日本微生物生態学会講演要旨集 25th YOSHI73 2009年 [査読無し][通常論文]
  • Yutaka Yawata, Hideaki Maseda, Satoshi Okabe, Akinobu Ito, Isao Sawada, Hiroaki Kurashima, Hiroo Uchiyama, Nobuhiko Nomura MICROBES AND ENVIRONMENTS 24 (4) 338 -342 2009年 [査読無し][通常論文]
     
    N-Octanoyl-L-homoserine lactone (C8-HSL) is an acyl-homoserine-lactone signal utilized in the quorum-sensing (QS) systems of Burkholderia cenocepacia and other bacterial species. Although also produced by Pseudomonas aeruginosa, its role in this species has not been elucidated. Here, we report that C8-HSL modulated antibiotic resistance and pyocyanin production in a P. aeruginosa efflux pump-deficient mutant. The rhl/las quorum-sensing system and qscR gene were both shown to be nonessential in the C8-HSL-induced changes in ofloxacin resistance, suggesting that P. aeruginosa possesses a distinct pathway to respond to C8-HSL.
  • SAVICHTCHEVA O, OKABE S Water Science and Technology 59 (9) 1831 -1840 2009年 [査読無し][通常論文]
     
    Rapid and reliable determination of the non-point sources of fecal pollution is a critical issue for the environmental microbiologists all over the world. In this work we evaluated the use of anaerobic bacterial group Bacteroides-Prevotella as an alternative fecal pollution indicator. Terminal restriction fragment length polymorphism (T-RFLP) and real-time polymerase chain reaction (RT-PCR) analyses were used to monitor and quantify human-, cow- and pig-specific fecal contamination in natural river waters. We also included DNA sequence analysis of the identified fecal markers revealed by T-RFLP in order to clarify the specificity of each marker. It was suggested that the most influent peaks for each fecal source could be used to identify the source of fecal pollution. Development of specific probes based on these markers permit to quantify source of contamination by quantitative RT-PCR. Therefore, we combined the T-RFLP results and RT-PCR assay to quantify fecal contamination by certain host. We can conclude that T-RFLP and RT-PCR analyses showed high reproducibility and sensitivity during analyzing real water samples and can be used to identify, track and quantify host-specific bacterial genetic markers in complex natural water environments. © IWA Publishing 2009.
  • Koji Kawata, Ryuhei Shimazaki, Satoshi Okabe ENVIRONMENTAL AND MOLECULAR MUTAGENESIS 50 (1) 46 -59 2009年01月 [査読無し][通常論文]
     
    Carcinogenesis is an important chronic toxicity of metals and metalloids, although their mechanisms of action are still unclear. Comparison of gene expression patterns induced by carcinogenic metals, metalloids, and model carcinogens would give an insight into understanding of their carcinogenic mechanisms. In this study, we examined the gene expression alteration in human hepatoma cell line, HepG2, after exposing to two metals (cadmium and nickel), a metalloid (arsenic), and three model carcinogenic chemicals N-dimethylnitrosoamine (DMN), 12-O-tetradeconoylphorbol-3-acetate (TPA), and tetrachloroethylene (TCE) using DNA microarrays with 8,795 human genes. Of the genes altered by As, Cd, and Ni exposures, 31-55% were overlapped with those altered by three model carcinogenic chemical exposures in our experiments. In particular, the metals and metalloid shared certain characteristics with TPA and TCE in remarkable upregulations of the genes associated with progression of cell cycle, which might play a central role in As, Cd, and Ni carcinogenesis. This characteristic of gene expression alteration was partially counteracted by intracellular accumulation of vitamin C in As-exposed cells, whereas the number of cell-cycle associated genes was increased in Cd- and Ni-exposed cells. In our experimental conditions, ROS might have an accelerative effect on the cell proliferation mechanisms of As, but have an inhibitory effect on those of other two heavy metals. Furthermore, based on the results of Q-PCR, the oncogene PTTG I, which was upregulated by all carcinogenic chemical exposures in the array experiments, might be a useful biomarker for evaluation of the carcinogenesis of inorganic carcinogens. Environ. Mol. Mutagen. 50:46-59, 2009. (C) 2008 Wiley-Liss, Inc.
  • 羽深昭, 坂井田健司, 佐藤久, 深澤達矢, 高橋正宏, 岡部聡 環境工学研究フォーラム講演集 45th 61 -63 2008年11月28日 [査読無し][通常論文]
  • 佐藤久, 池田真之, 高橋慶多, 深澤達矢, 高橋正宏, 岡部聡, 中原禎仁, 笹川学 環境工学研究フォーラム講演集 45th 58 -60 2008年11月28日 [査読無し][通常論文]
  • Thithiwat May, Satoshi Okabe JOURNAL OF BACTERIOLOGY 190 (22) 7479 -7490 2008年11月 [査読無し][通常論文]
     
    It has been shown that Escherichia coli harboring the derepressed IncFI and IncFII conjugative F plasmids form complex mature biofilms by using their F-pilus connections, whereas a plasmid-free strain forms only patchy biofilms. Therefore, in this study we investigated the contribution of a natural IncF conjugative F plasmid to the formation of E. coli biofilms. Unlike the presence of a derepressed F plasmid, the presence of a natural IncF F plasmid promoted biofilm formation by generating the cell-to-cell mating F pili between pairs of F(+) cells (approximately two to four pili per cell) and by stimulating the formation of colanic acid and curli meshwork. Formation of colanic acid and curli was required after the initial deposition of F-pilus connections to generate a three-dimensional mushroom-type biofilm. In addition, we demonstrated that the conjugative factor of F plasmid, rather than a pilus synthesis function, was involved in curli production during biofilm formation, which promoted cell-surface interactions. Curli played an important role in the maturation process. Microarray experiments were performed to identify the genes involved in curli biosynthesis and regulation. The results suggested that a natural F plasmid was more likely an external activator that indirectly promoted curli production via bacterial regulatory systems (the EnvZ/OmpR two-component regulators and the RpoS and HN-S global regulators). These data provided new insights into the role of a natural F plasmid during the development of E. coli biofilms.
  • Yuki Miura, Satoshi Okabe ENVIRONMENTAL SCIENCE & TECHNOLOGY 42 (19) 7380 -7386 2008年10月 [査読無し][通常論文]
     
    We, for the first time, quantitatively determined cell specific uptake activities of microbial products (bacterial cell detritus and extracellular polymeric substances, EPS) by the member of uncultured Chloroflexi by using a microautoradiography combined with fluorescence in situ hybridization (MAR-FISH) technique. For this MAR-FISH analysis, we prepared [(14)C]-labeled microbial products from biomass sludge obtained and bacterial strains (Pseudomonas sp. and Acinetobacter sp.) isolated from our pilot-scale membrane bioreactor (MBR) as tracer substrates, which probably represent the more realistic food source in the MBR. The quantitative MAR-FISH analyses clearly showed that most of the uncultured Chloroflexi could indeed uptake the bacterial detritus of the two isolated strains with rates of 1.7-3.5 x 10(-17) g-C mu m(-2)-surface area h(-1) (corresponding to 1.2-1.7 mg-C-bacterial detritus L(-1) h(-1)) in the cultures, which were, however, about 2 orders of magnitude lower than the uptake rates of simple monosaccharides (mannose, arabinose, fucose, and galactose). Based on these results and their high abundance (more than 20% of total bacteria detected with EUB338-mixed probes), it could be estimated that the uncultured Chloroflexi contributes 38-51% of the total degradation of microbial products occurred in the MAR-FISH cultures.
  • 佐藤久, 高橋慶多, 深澤達矢, 高橋正宏, 岡部聡, 中原禎仁, 笹川学 土木学会年次学術講演会講演概要集(CD-ROM) 63rd (Disk 2) ROMBUNNO.7-043 2008年08月13日 [査読無し][通常論文]
  • S. -J. Cho, S. Okabe, S. -J. Lee ENVIRONMENTAL TECHNOLOGY 29 (4) 463 -471 2008年04月 [査読無し][通常論文]
     
    To investigate the inorganic nitrogen conversion in reactors that were operated under aerobic and micro-aerobic conditions and to identify populations that became acclimated in the reactors under those oxygen conditions, we operated two reactors with 72 h of hydraulic retention time and an artificial medium containing ammonium- and nitrite-nitrogen without any organic compound. We determined the concentration of inorganic nitrogen in samples from the reactors and the microbial community structure in the reactors by using PCR-DGGE of the partial 16S rRNA gene. The results showed that nitrification to nitrate fully progressed in the aerobic reactor within 24 h, but we could not find any obvious reaction in the micro-aerobic reactor. In a view of microbial community structure, the total number of microorganisms composing the communities in the reactors was dramatically decreased compared to those of the initial inoculated sludge that originated from the Jang-lim, sewage treatment plant in Busan, Republic of Korea because of the limited nutrients present. One Nitrospira sp. was clearly detected under both aerobic (as DO > 2 mg l(-1)) and micro-aerobic (as DO < 0.7 mg l(-1)) conditions while no AOB-Iike bacterium was exactly matched among main bands. By the results, nitrite-oxidizing bacteria may be more tolerant to variations in oxygen, or the occupation of ammonia-oxidizing bacteria as dominant groups may have been inhibited by substrate starvation and high concentrations of nitrite-N in influent.
  • Akinobu Ito, Thithiwat May, Koji Kawata, Satoshi Okabe BIOTECHNOLOGY AND BIOENGINEERING 99 (6) 1462 -1471 2008年04月 [査読無し][通常論文]
     
    Presence of starved, stationary phase-like zones in biofilms seems to be an important factor for biofilm formation. In this study, roles of rpoS gene in the formation of Escherichia coli biofilms were investigated. E. coli MG 1655 wild type (WT) and rpoS mutant (Delta rpoS) strains were used to compare biofilm formation capacity and global gene expression. Even though the Delta rpoS strain could attach and form microcolonies on glass surfaces, it could not establish mature biofilms. DNA microarray analysis revealed that WT biofilms (WBF) showed similar pattern of gene expression with WT planktonic stationary phase, whereas Delta rpoS biofilms (MBF) showed similar pattern of gene expression with WT planktonic exponential phase. Genes involved in energy metabolism (arpIBEFHAG, atpC, cydAB) and flagella synthesis (flgB, flgC, flhD, fliA, fliC, fliY) showed increased expression in the MBF, but not in the WBF. Moreover, genes involved in stress responses (blc, cspG, dinD poxB, wcaF, wcaI, and yfcf) showed increased expression in the WBF compared to the MBF. These results suggested that the rpoS gene contributed in maturation of E. coli biofilms through regulation of global gene expression including energy metabolism, motility, and stress responses.
  • 三浦佑己, 渡辺義公, 岡部聡 日本水環境学会年会講演集 42nd 170 2008年03月19日 [査読無し][通常論文]
  • 佐藤久, 坂井田健司, 岡部聡, 渡辺義公 環境バイオテクノロジー学会大会プログラム講演要旨集 34th 27 2008年 [査読無し][通常論文]
  • 佐藤久, 坂井田健司, 岡部聡, 渡辺義公 環境工学研究フォーラム講演集 44th 224 -226 2007年11月16日 [査読無し][通常論文]
  • Hisashi Satoh, Yuki Miura, Ikuo Tsushima, Satoshi Okabe APPLIED AND ENVIRONMENTAL MICROBIOLOGY 73 (22) 7300 -7307 2007年11月 [査読無し][通常論文]
     
    The microbial community structure and spatial distribution of microorganisms and their in situ activities in anaerobic granules were investigated by 16S rRNA gene-based molecular techniques and microsensors for CH4, H-2, pH, and the oxidation-reduction potential (ORP). The 16S rRNA gene-cloning analysis revealed that the clones related to the phyla Alphaproteobacteria (detection frequency, 51%), Firmicutes (20%), Chloroflexi (9%), and Betaproteobacteria (8%) dominated the bacterial clone library, and the predominant clones in the archaeall clone library were affiliated with Methanosaeta (73%). In situ hybridization with oligonucleotide probes at the phylum level revealed that these microorganisms were numerically abundant in the granule. A layered structure of microorganisms was found in the granule, where Chloroflexi and Betaproteobacteria were present in the outer shell of the granule, Firmicutes were found in the middle layer, and aceticlastic Archaea were restricted to the inner layer. Microsensor measurements for CH4, H-2, pH, and ORP revealed that acid and H-2, production occurred in the upper part of the granule, below which H-2, consumption and CH4 production were detected. Direct comparison of the in situ activity distribution with the spatial distribution of the microorganisms implied that Chloroflexi contributed to the degradation of complex organic compounds in the outermost layer, H-2 was produced mainly by Firmicutes in the middle layer, and Methanosaeta produced CH4 in the inner layer. We determined the effective diffusion coefficient for H-2 in the anaerobic granules to be 2.66 X 10(-5) cm(2) s(-1), which was 57% in water.
  • Yuki Miura, Yoshimasa Watanabe, Satoshi Okabe ENVIRONMENTAL SCIENCE & TECHNOLOGY 41 (22) 7787 -7794 2007年11月 [査読無し][通常論文]
     
    We operated pilot-scale submerged membrane bioreactors (MBR) treating real municipal wastewater for over 3 months and observed an interesting phenomenon that carbohydrate concentrations in the MBRs rapidly increased, which consequently resulted in membrane fouling, when relative abundance of the member of uncultured Chloroflexi decreased from over 30% of total Bacteria to less than 10%. We, therefore, hypothesized that the uncultured Chloroflexi present in the MBRs could preferentially degrade carbohydrates and consequently prevent membrane fouling. To test this hypothesis, we investigated the phylogenetic identity, diversity, and in situ physiology (substrate utilization characteristics) of Chloroflexi residing in the MBR by using 16S rRNA gene sequencing analysis and microautoradiography combined with fluorescence in situ hybridization (MAR-FISH) technique. Most of the clones related to the phylum Chloroflexi were affiliated with the Chloroflexi subphylum 1 containing only a few cultured representatives. The MAR-FISH revealed that the members of Chloroflexi were metabolically versatile and could preferentially utilize glucose and N-acetyl glucosamine (a main substantial constituent of the cell wall peptidoglycan) under oxic and anoxic conditions. The utilization of these compounds was low at low pH. These findings suggest that the members of Chloroflexi are ecologically significant in the MBR treating municipal wastewater and are responsible for degradation of SMP including carbohydrates and cellular materials, which consequently reduces membrane fouling potential.
  • Satoshi Okabe, Yoko Shimazu APPLIED MICROBIOLOGY AND BIOTECHNOLOGY 76 (4) 935 -944 2007年09月 [査読無し][通常論文]
     
    Host-specific Bacteroides-Prevotella 16S rRNA genetic markers are promising alternative indicators for identifying the sources of fecal pollution because of their high abundance in the feces of warm-blooded animals and high host specificity. However, little is known about the persistence of these genetic markers in environments after being released into environmental waters. The persistence of feces-derived four different host-specific Bacteroides-Prevotella 16S rRNA genetic makers (total, human-, cow-, and pig-specific) in environmental waters was therefore investigated at different incubation temperatures (4, 10, 20, and 30 degrees C) and salinities (0, 10, 20, and 30 ppt) and then compared with the survival of conventional fecal-indicator organisms. The host-specific genetic markers were monitored by using real-time polymerase chain reaction (PCR) assays with specific primer sets. Each host-specific genetic marker showed similar responses in non-filtered river water and seawater: They persisted longer at lower temperatures and higher salinities. In addition, these markers did not increase in all conditions tested. Decay rates for indicator organisms were lower than those for host-specific genetic markers at temperature above 10 degrees C. Furthermore, we investigated whether the PCR-detectable 16S rRNA genetic markers reflect the presence of live target cells or dead target cells in environmental waters. The result revealed that the detection of the Bacteroides-Prevotella 16S rRNA genetic markers in environmental waters mainly reflected the presence of 'viable but non-culturable' Bacteroides-Prevotella cells. These findings indicate that seasonal and geographical variations in persistence of these host-specific Bacteroides-Prevotella 16S rRNA genetic markers must be considered when we use them as alternative fecal indicators in environmental waters.
  • Tomonori Kindaichi, Ikuo Tsushima, Yuji Ogasawara, Masaki Shimokawa, Noriatsu Ozaki, Hisashi Satoh, Satoshi Okabe APPLIED AND ENVIRONMENTAL MICROBIOLOGY 73 (15) 4931 -4939 2007年08月 [査読無し][通常論文]
     
    We investigated autotrophic anaerobic ammonium-oxidizing (anammox) biofilms for their spatial organization, community composition, and in situ activities by using molecular biological techniques combined with microelectrodes. Results of phyllogenetic analysis and fluorescence in situ hybridization (FISH) revealed that "Brocadia"-like anammox bacteria that hybridized with the Amx820 probe dominated, with 60 to 92% of total bacteria in the upper part (< 1,000 mu m) of the biofilm, where high anammox activity was mainly detected with microelectrodes. The relative abundance of anammox bacteria decreased along the How direction of the reactor. FISH results also indicated that Nitrosomonas-, Nitrosospira-, and Nitrosococcus-like aerobic ammonia-oxidizing bacteria (AOB) and Nitrospira-like nitrite-oxidizing bacteria (NOB) coexisted with anammox bacteria and accounted for 13 to 21% of total bacteria in the biofilms. Microelectrode measurements at three points along the anammox reactor revealed that the NH4+, and NO2- consumption rates decreased from 0.68 and 0.64 mu mol cm(-2) h(-1) at P2 (the second port, 170 mm from the inlet port) to 0.30 and 0.35 mu mol cm(-2) h(-1) at P3 (the third port, 205 rum from the inlet port), respectively. No anammox activity was detected at P4 (the fourth port, 240 mm from the inlet port), even though sufficient amounts of NH4+ and NO2- and a high abundance of anammox bacteria were still present. This result could be explained by the inhibitory effect of organic compounds derived from biomass decay and/or produced by anammox and coexisting bacteria in the upper parts of the biofilm and in the upstream part of the reactor. The anammox activities in the biofilm determined by microelectrodes reflected the overall reactor performance. The several groups of aerobic AOB lineages, Nitrospira-like NOB, and Betaproteobacteria coexisting in the anammox biofilm might consume a trace amount of O-2, or organic compounds, which consequently established suitable microenvironments for anammox bacteria.
  • Olga Savichtcheva, Noriko Okayama, Satoshi Okabe WATER RESEARCH 41 (16) 3615 -3628 2007年08月 [査読無し][通常論文]
     
    Occurrence and prevalence of different bacterial enteric pathogens as well as their relationships with conventional (total and fecal coliforms) and alternative fecal indicators (host-specific Bacteroides 16S rRNA genetic markers) were investigated for various water samples taken from different sites with different degrees of fecal contamination. The results showed that a wide range of bacterial pathogens could be detected in both municipal wastewater treatment plant samples and in surface water samples. Logistic regression analysis revealed that total and human-specific Bacteroides 16S rRNA genetic markers showed significant predictive values for the presence of Escheriachia coli O-157, Salmonella, heat-labile enterotoxin (IT) of enterotoxigenic E. coli (ETEC), and heat-stable enterotoxin for human (STh) of ETEC. The probability of occurrence of these pathogenic bacteria became significantly high when the concentrations of human-specific and total Bacteroides 16S rRNA genetic markers exceeded 10(3) and 104 copies/100mL. In contrast, Clostridium perfringens was detected at high frequency regardless of sampling sites and levels of Bacteroides 16S rRNA genetic markers. No genes related to Shigella spp., Staphylococcus aureus and Vibrio cholerae were detected in any samples analyzed in this study. Conventional indicator microorganisms had low levels of correlation with the presence of pathogens as compared with the alternative fecal indicators. These results suggested that real-time PCR-based measurement of alternative Bacteroides 16S rRNA genetic markers was a rapid and sensitive tool to identify host-specific fecal pollution and probably associated bacterial pathogens. However, since one fecal indicator might not represent the relative abundance of all pathogenic bacteria, viruses and protozoa, combined application of alternative indicators with conventional ones could provide more comprehensive pictures of fecal contamination, its source and association with pathogenic microorganisms. (c) 2007 Elsevier Ltd. All rights reserved.
  • Herto Dwi Ariesyady, Tsukasa Ito, Kazurni Yoshiguchi, Satoshi Okabe APPLIED MICROBIOLOGY AND BIOTECHNOLOGY 75 (3) 673 -683 2007年06月 [査読無し][通常論文]
     
    The phylogenetic and functional diversity of syntrophic prop ionate-oxidizing bacteria (POB) present in an anaerobic digester was investigated by microautoradiography combined with fluorescent in situ hybridization (MAR-FISH) that can directly link 16S rRNA phylogeny with in situ metabolic function. The syntrophic POB community in the anaerobic digester sludge consisted of at least four phylogenetic groups: Syntrophobacter, uncultured short rod Smithella (Smithella sp. SR), uncultured long rod Smithella (Smithella sp. LR), and an unidentified group. The activities of these POB groups were dependent on the propionate concentrations. The uncultured Smithella sp. SR accounted for 52-62% of the total active POB under all the propionate concentrations tested (0.515 mM). In contrast, uncultured Smithella sp. LR was active only at lower propionate concentrations and became a dominant active POB at 0.5 mM of propionate. Syntrophobactet- accounted for 16-3 1 % of the total active POB above 2.5 mM propionate, whereas the active Syntrophobacter population became low (ca. 6%) at 0.5 mM of propionate. The anaerobic digester was operated in a fill and draw mode, resulting in periodical changes in propionate concentration ranging from 0 to 10 mM. These phylogenetically and functionally diverse, to some extent functionally redundant, active POB communities were dynamically responding to the periodical changes in propionate concentration.
  • Koji Kawata, Hiroyuki Yokoo, Ryuhes Shimazaki, Satoshi Okabe ENVIRONMENTAL SCIENCE & TECHNOLOGY 41 (10) 3769 -3774 2007年05月 [査読無し][通常論文]
     
    Microarray technology is proving to be a useful tool to classify undefined environmental toxicants, to investigate underlying mechanisms of toxicity, and to identify candidate toxicant-specific genetic markers by examining global effects of putative toxicants on gene expression profiles. The aim of this study was to evaluate the toxicities of six heavy metals through the comparison with gene expression patterns induced by well-known chemicals. For this purpose, we first identified the genes altered specifically in HepG2 under the exposure of 2,3-dimethoxy-1,4-naphthoquinone (DMNQ), phenol, and N-nitrosodimethylamine (DMN), which were selected as the model chemicals, using DNA microarray. On the basis of the expression profiles of these genes, toxicities of six heavy metals, arsenic, cadmium, nickel, antimony, mercury, and chromium, were evaluated. The specific gene alteration and hierarchical clustering revealed that biological action of six heavy metals was clearly related to that of DMNQ which has been reported to be a reactive oxygen species (ROS) generating chemical and which induced the genes associated with cell proliferative responses. These results suggest that cell proliferative responses which are probably caused by ROS are a major apparent biological action of high-dose heavy metals, supporting the previous reports. Overall, a mechanism-based classification by DNA microarray would be an efficient method for evaluation of toxicities of environmental samples.
  • Ikuo Tsushima, Yuji Ogasawara, Tomonori Kindaichi, Hisashi Satoh, Satoshi Okabe WATER RESEARCH 41 (8) 1623 -1634 2007年04月 [査読無し][通常論文]
     
    To promptly establish anaerobic ammonium oxidation (anammox) reactors, appropriate seeding sludge with high abundance and activity of anammox bacteria was selected by quantifying 16S rRNA gene copy numbers of anammox bacteria by real-time quantitative PCR (RTQ-PCR) and batch culture experiments. The selected sludge was then inoculated into up-flow fixed-bed biofilm column reactors with nonwoven fabric sheets as biomass carrier and the reactor performances were monitored over 1 year. The anammox reaction was observed within 50 days and a total nitrogen removal rate of 26.0 kg-N m(-3) day(-1) was obtained after 247 days. To our knowledge, such a high rate has never been reported before. Hydraulic retention time (HRT) and influent NH4+ to NO2- molar ratio could be important determinant factors for efficient nitrogen removal in this study. The higher nitrogen removal rate was obtained at the shorter HRT and higher influent NH4+/NO2- molar ratio. After anammox reactors were fully developed, the community structure, spatial organization and in situ activity of the anammox biofilms were analyzed by the combined use of a full-cycle of 16S rRNA approach and microelectrodes. In situ hybridization results revealed that the probe Amx820-hybridized anaerobic anammox bacteria were distributed throughout the biofilm (accounting for more than 70% of total bacteria), They were associated with Nitrosomonas-like aerobic ammonia-oxidizing bacteria (AAOB) in the surface biofilm. The anammox bacteria present in this study were distantly related to the Candidatus Brocadia anammoxidans with the sequence similarity of 95%. Microelectrode measurements showed that a high in situ anammox activity (i.e., simultaneous consumption of NH4+ and NO2-) of 4.45 g-N of (NH4++NO2-) m(-2) day(-1) was detected in the upper 800 mu m of the biofilm, which was consistent with the spatial distribution of anammox bacteria. (c) 2007 Elsevier Ltd. All rights reserved.
  • Herto Dwi Ariesyady, Tsukasa Ito, Satoshi Okabe Water research 41 (7) 1554 -68 2007年04月 [査読有り][通常論文]
     
    Functional Bacteria and Archaea community structures of a full-scale anaerobic sludge digester were investigated by using a full-cycle 16S rRNA approach followed by microautoradiography (MAR)-fluorescent in situ hybridization (FISH) technique and micromanipulation. FISH analysis with a comprehensive set of 16S and 23S rRNA-targeted oligonucleotide probes based on 16S rRNA clone libraries revealed that the Gram-positive bacteria represented by probe HGC69A-hybridized Actinobacteria (8.5+/-1.4% of total 4', 6-diamidino-2-phenylindole (DAPI)-stained cells) and probe LGC354-hybridized Firmicutes (3.8+/-0.8%) were the major phylogenetic bacterial phyla, followed by Bacteroidetes (4.0+/-1.2%) and Chloroflexi (3.7+/-0.8%). The probe MX825-hybridized Methanosaeta (7.6+/-0.8%) was the most abundant archaeal group, followed by Methanomicrobiales (2.8+/-0.6%) and Methanobacteriaceae (2.7+/-0.4%). The functional community structures (diversity and relative abundance) of major trophic groups were quantitatively analyzed by MAR-FISH. The results revealed that glucose-degrading microbial community had higher abundance (ca. 10.6+/-4.9% of total DAPI-stained cells) and diversity (at least seven phylogenetic groups) as compared with fatty acid-utilizing microbial communities, which were more specialized to a few phylogenetic groups. Despite the dominance of Betaproteobacteria, members of Chloroflexi, Smithella, Syntrophomonas and Methanosaeta groups dominated the [(14)C]glucose-, [(14)C]propionate-, [(14)C]butyrate- and [(14)C]acetate-utilizing microorganism community, and accounted for 27.7+/-4.3%, 29.6+/-7.0%, 34.5+/-7.6% and 18.2+/-9.5%, respectively. In spite of low abundance (ca. 1%), the hitherto unknown metabolic functions of Spirochaeta and candidate phylum of TM7 as well as Synergistes were found to be glucose and acetate utilization, respectively.
  • 坂井田健司, 佐藤久, 岡部聡, 渡辺義公 日本水環境学会年会講演集 41st 335 2007年03月15日 [査読無し][通常論文]
  • 三浦佑己, 渡辺義公, 岡部聡 日本水環境学会年会講演集 41st 227 2007年03月15日 [査読無し][通常論文]
  • Satoshi Okabe, Noriko Okayama, Olga Savichtcheva, Tsukasa Ito APPLIED MICROBIOLOGY AND BIOTECHNOLOGY 74 (4) 890 -901 2007年03月 [査読無し][通常論文]
     
    Based on the comparative 16S rRNA gene sequence analysis of fecal DNAs, we identified one human-, three cow-, and two pig-specific Bacteroides-Prevotella 16S rRNA genetic markers, designed host-specific real-time polymerase chain reaction (real-time PCR) primer sets, and successfully developed real-time PCR assay to quantify the fecal contamination derived from human, cow, and pig in natural river samples. The specificity of each newly designed host-specific primer pair was evaluated on fecal DNAs extracted from these host feces. All three cow-specific and two pig-specific primer sets amplified only target fecal DNAs (in the orders of 9-11 log(10) copies per gram of wet feces), showing high host specificity. This real-time PCR assay was then applied to the river water samples with different fecal contamination sources and levels. It was confirmed that this assay could sufficiently discriminate and quantify human, cow, and pig fecal contamination. There was a moderate level of correlation between the Bacteroides-Prevotella group-specific 16S rRNA gene markers with fecal coliforms (r(2) = 0.49), whereas no significant correlation was found between the human-specific Bacteroides 16S rRNA gene with total and fecal coliforms. Using a simple filtration method, the minimum detection limits of this assay were in the range of 50-800 copies/100 ml. With a combined sample processing and analysis time of less than 8 h, this real-time PCR assay is useful for monitoring or identifying spatial and temporal distributions of host-specific fecal contaminations in natural water environments.
  • Yuki Miura, Mirian Noriko Hiraiwa, Tsukasa Ito, Takanori Itonaga, Yoshimasa Watanabe, Satoshi Okabe WATER RESEARCH 41 (3) 627 -637 2007年02月 [査読無し][通常論文]
     
    Bacterial community structures in pilot-scale conventional membrane bioreactors (CMBRs) and hybrid MBRs (HMBRs) which were combined with pre-coagulation/sedimentation were analyzed by polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) and fluorescence in situ hybridization (FISH) techniques. The results were compared with the community structure in a full-scale activated sludge (AS) process treating the same municipal wastewater. The Dice index (Cs) of similarity analysis of DGGE banding patterns demonstrated that the microbial community in AS was more similar to those in CMBR1 and CMBR2 than HMBR1 and HMBR2. This suggested that influent wastewater composition had a larger impact on bacterial community structures. Long-term community structure changes in the HMBRs and CMBRs were monitored and analyzed over 240 days by Non-metric multidimensional scaling (NMDS) analysis of DGGE banding patterns. The NMDS analysis revealed that both HMBRs and CMBRs had marked changes in community structures during the first about 100 days. Thereafter the perpetual fluctuations of bacterial community structures were observed in both HMBRs and CMBRs, even though the stable MBR performances (the performance was measured as membrane permeability and removal of dissolved organic carbon, DOC) were achieved. These results suggest that not only the stability, but also the adequate dynamics ("flexibility") of the bacterial community structure are important for the stable performance of the MBRs treating complex municipal wastewater. (c) 2006 Elsevier Ltd. All rights reserved.
  • Ikuo Tsushima, Tomonori Kindaichi, Satoshi Okabe WATER RESEARCH 41 (4) 785 -794 2007年02月 [査読無し][通常論文]
     
    The anaerobic ammonium-oxidizing (ANAMMOX) bacteria were enriched from a rotating disk reactor (RDR) biofilm in semi-batch cultures. Based on fluorescence in situ hybridization (FISH) analysis, this enrichment led to a relative population size of 36% ANAMMOX bacteria. Phylogenetic analysis revealed that all the detected clones were related to the previously reported ANAMMOX bacteria, Candidatus Brocadia anammoxidans (AF375994), with 92% sequence similarity. Furthermore, we successfully developed a real-time polymerase chain reaction (PCR) assay to quantify populations of ANAMMOX bacteria in the enrichment cultures. For this real-time PCR assay, PCR primer sets targeting 16S ribosomal RNA genes of ANAMMOX bacteria were designed and used. The quantification range of this assay was 6 orders of magnitude, from 8.9 x 10(1) to 8.9 x 10(6) copies per PCR, corresponding to the detection limit of 3.6 x 10(3) target copies mL(-1). A significant correlation was found between the increase in copy numbers of 16S rRNA gene of ANAMMOX bacteria and the increase in nitrogen removal rates in the enrichment cultures. Quantifying ANAMMOX bacterial populations in the enrichment culture made it possible to estimate the doubling time of the enriched ANAMMOX bacteria to be 3.6 to 5.4 days. The real-time PCR assay gave comparable population sizes in the enrichment cultures with the FISH results. These results suggest that the real-time PCR assay developed in this study is useful and reliable for quantifying the populations of ANAMMOX bacteria in environmental and engineering samples. (c) 2006 Elsevier Ltd. All rights reserved.
  • Hisashi Satoh, Yoshiyuki Nakamura, Satoshi Okabe APPLIED AND ENVIRONMENTAL MICROBIOLOGY 73 (4) 1341 -1348 2007年02月 [査読無し][通常論文]
     
    Influences of infaunal burrows constructed by the polychaete (Tylorrhynchus heterochaetus) on O-2 concentrations and community structures and abundances of ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB) in intertidal sediments were analyzed by the combined use of a 16S rRNA gene-based molecular approach and microelectrodes. The microelectrode measurements performed in an experimental system developed in an aquarium showed direct evidence of O-2 transport down to a depth of 350 mm of the sediment through a burrow. The 16S rRNA gene-cloning analysis revealed that the betaproteobacterial AOB communities in the sediment surface and the burrow walls were dominated by Nitrosomonas sp. strain Nm143-like sequences, and most of the clones in Nitrospira-like NOB clone libraries of the sediment surface and the burrow walls were related to the Nitrospira marina lineage. Furthermore, we investigated vertical distributions of AOB and NOB in the infaunal burrow walls and the bulk sediments by real-time quantitative PCR (Q-PCR) assay. The AOB and Nitrospira-like NOB-specific 16S rRNA gene copy numbers in the burrow walls were comparable with those in the sediment surfaces. These numbers in the burrow wall at a depth of 50 to 55 rum from the surface were, however, higher than those in the bulk sediment at the same depth. The microelectrode measurements showed higher NH4+ consumption activity at the burrow wall than those at the surrounding sediment. This result was consistent with the results of microcosm experiments showing that the consumption rates of NH4+ and total inorganic nitrogen increased with increasing infaunal density in the sediment. These results clearly demonstrated that the infaunal burrows stimulated O-2 transport into the sediment in which otherwise reducing conditions prevailed, resulting in development of high NH4+ consumption capacity. Consequently, the infaunal burrow became an important site for NH4+ consumption in the intertidal sediment.
  • Satoshi Okabe, Mitsunori Odagiri, Tsukasa Ito, Hisashi Satoh APPLIED AND ENVIRONMENTAL MICROBIOLOGY 73 (3) 971 -980 2007年02月 [査読無し][通常論文]
     
    Microbially induced concrete corrosion (MICC) in sewer systems has been a serious problem for a long time. A better understanding of the succession of microbial community members responsible for the production of sulfuric acid is essential for the efficient control of MICC. In this study, the succession of sulfur-oxidizing bacteria (SOB) in the bacterial community on corroding concrete in a sewer system in situ was investigated over I year by culture-independent 16S rRNA gene-based molecular techniques. Results revealed that at least six phylotypes of SOB species were involved in the MICC process, and the predominant SOB species shifted in the following order: Thiothrix sp., Thiobacillus plumbophilus, Thiomonas intermedia, Halothiobacillus neapolitanus, Acidiphilium acidophilum, and Acidithiobacillus thiooxidans. A. thiooxidans, a hyperacidophilic SOB, was the most dominant (accounting for 70% of EUB338-mixed probe-hybridized cells) in the heavily corroded concrete after I year. This succession of SOB species could be dependent on the pH of the concrete surface as well as on trophic properties (e.g., autotrophic or mixotrophic) and on the ability of the SOB to utilize different sulfur compounds (e.g., H2S, S-0, and S2O32-). In addition, diverse heterotrophic bacterial species (e.g., halo-tolerant, neutrophilic, and acidophilic bacteria) were associated with these SOB. The microbial succession of these microorganisms was involved in the colonization of the concrete and the production of sulfuric acid. Furthermore, the vertical distribution of microbial community members revealed that A. thiooxidans was the most dominant throughout the heavily corroded concrete (gypsum) layer and thatA. thiooxidans was most abundant at the highest surface (1.5-mm) layer and decreased logarithmically with depth because of oxygen and H2S transport limitations. This suggested that the production of sulfuric acid by A. thiooxidans occurred mainly on the concrete surface and the sulfuric acid produced penetrated through the corroded concrete layer and reacted with the sound concrete below.
  • Satoshi Okabe Sulphate-Reducing Bacteria: Environmental and Engineered Systems 359 -382 2007年01月01日 [査読無し][通常論文]
     
    INTRODUCTION Sulphate-reducing bacteria (SRB) are a phylogenetically and physiologically diverse group of bacteria, characterized by their versatile metabolic capability to use various electron acceptors and donors (Widdel, 1988). SRB are therefore universally distributed in diverse environments and play significant ecophysiological roles in anaerobic biomineralization pathways. The degradation of organic matter by a complex microbial community is governed to a large extent by available electron acceptors. The terminal stages of the anaerobic mineralization of organic matter is catalyzed by SRB and methanogens, and their competitive and cooperative interactions have been described previously (Oude Elferink et al., 1994). Typical domestic wastewaters contain sulphate concentrations of 100-1000 µM and relatively low dissolved oxygen due to the lower solubility and rapid depletion of this gas by biological activity. Thus, sulphate reduction can be the dominant terminal electron accepting process and account for up to 50% of mineralization of organic matter in wastewater biofilms (Kühl and Jorgensen, 1992 Okabe et al., 2003a). Multiple electron donors and electron acceptors are present in the wastewaters. As a result, wastewater biofilms are very complex multispecies biofilms, displaying considerable heterogeneity, with regard to both the microorganisms present and their physicochemical microenvironments. Sulphate reduction is anticipated to take place in the deeper anoxic biofilm strata even though the bulk liquid is oxygenated. It is, therefore, thought that a successive vertical zonation of respiratory processes can be found in aerobic wastewater biofilms with a typical thickness of only a few millimeters (Ito et al., 2002b Kühl and Jorgensen, 1992 Okabe et al., 1999a 2003a Ramsing et al., 1993).
  • 環境工学論文集 44 201 -206 2007年 [査読無し][通常論文]
  • Ikuo Tsushima, Yuji Ogasawara, Masaki Shimokawa, Tomonori Kindaichi, Satoshi Okabe WATER SCIENCE AND TECHNOLOGY 55 (8-9) 9 -17 2007年 [査読無し][通常論文]
     
    The anaerobic ammonium oxidation (Anammox) process is a new efficient and cost effective method of ammonium removal from wastewater. Under strictly anoxic condition, ammonium is directly oxidised with nitrite as electron acceptor to dinitrogen gas. However, it is extremely difficult to cultivate Anammox bacteria due to their low growth rate. This suggests that a rapid and efficient start-up of Anammox process is the key to practical applications. To screen appropriate seeding sludge with high Anammox potential, a real-time quantitative PCR assay with newly designed primers has been developed. Thereafter, the seeding sludge with high abundance of Anammox bacteria (1.7 X 10(8) copies/mg-dry weight) was selected and inoculated into an upflow anaerobic biofilters (UABs). The UABs were operated for more than 1 year and the highest nitrogen removal rate of 24.0 kg(-N) m(-3) day(-1) was attained. In addition, the ecophysiology of Anammox bacteria (spatial distribution and in situ activity) in biofilms was analysed by combining a full-cycle 16S rRNA approach and microelectrodes. The microelectrode measurement clearly revealed that a successive vertical zonation of the partial nitrification (NH4+ to NO2-), Anammox reaction and denitrification was developed in the biofilm in the UAB. This result agreed with the spatial distribution of corresponding bacterial populations in the biofilm. We linked the micro-scale information (i.e. single cell and/or biofilm levels) with the macro-scale information (i.e. the reactor level) to understand the details of Anammox reaction occurring in the UABs.
  • Yuki Miura, Yoshimasa Watanbe, Satoshi Okabe ENVIRONMENTAL SCIENCE & TECHNOLOGY 41 (2) 632 -638 2007年01月 [査読無し][通常論文]
     
    For more efficient control and prediction of membrane biofouling in membrane bioreactors (MBRs), a fundamental understanding of mechanisms of membrane biofouling is essential. In this study, we operated full-scale submerged MBRs using real municipal wastewater delivered from the primary sedimentation basin of a municipal wastewater treatment facility over 3 months, and the adhesion and formation of biofilms on 0.4-mu m pore size polyethylene hollow-fiber microfiltration (MF) membrane surfaces, separated from simple deposition of sludge cake, were monitored using scanning electron microscopy (SEM). In addition, the compositions of planktonic and biofilm microbial communities in the MBR were analyzed using culture independent molecular-based methods (i.e., fluorescent in situ hybridization (FISH) and 16S rRNA gene sequence analysis). The SEM and LIVE/DEAD staining analyses clearly showed that the biofilms gradually developed on the membrane surfaces with time, which had a strong positive correlation with the increase in trans-membrane pressure (TMP). This indicated that the biofilm formation induced the membrane fouling. The FISH results revealed that the microbial communities on membrane surfaces were quite different from those in the planktonic biomass in the mixed liquor. Moreover, FISH and 16S rRNA gene sequence analyses revealed that a specific phylogenetic group of bacteria, the Betaproteobacteria, probably played a major role in development of the mature biofilms, which led to the severe irreversible membrane biofouling.
  • Tomonori Kindaichi, Yoshiko Kawano, Tsukasa Ito, Hisashi Satoh, Satoshi Okabe BIOTECHNOLOGY AND BIOENGINEERING 94 (6) 1111 -1121 2006年08月 [査読無し][通常論文]
     
    Population dynamics of ammonia-oxidizing bacteria (AOB) and uncultured Nitrospira-like nitrite-oxidizing bacteria (NOB) dominated in autotrophic nitrifying biofilms were determined by using real-time quantitative polymerase chain reaction (RTQ-PCR) and fluorescence in situ hybridization (FISH). Although two quantitative techniques gave the comparable results the RTQ-PCR assay was easier and faster than the FISH technique for quantification of both nitrifying bacteria in dense microcolony-forming nitrifying biofilms. Using this RTQ-PCR assay, we could successfully determine the maximum specific growth rate (mu = 0.021/h) of uncultured Nitrospira-like NOB in the suspended enrichment culture. The population dynamics of nitrifying bacteria in the biofilm revealed that once they formed the biofilm, the both nitrifying bacteria grew slower than in planktonic cultures. We also calculated the spatial distributions of average specific growth rates of both nitrifying bacteria in the biofilm based on the concentration profiles of NH4+, NO2-, and O-2, which were determined by microelectrodes, and the double-Monod model. This simple model estimation could explain the stratified spatial distribution of AOB and Nitrospira-like NOB in the biofilm. The combination of culture-independent molecular techniques and microelectrode measurements is a very powerful approach to analyze the in situ kinetics and ecophysiology of nitrifying bacteria including uncultured Nitrospira-like NOB in complex biofilm communities. (c) 2006 Wiley Periodicals, Inc.
  • H Satoh, T Yamakawa, T Kindaichi, T Ito, S Okabe BIOTECHNOLOGY AND BIOENGINEERING 94 (4) 762 -772 2006年07月 [査読無し][通常論文]
     
    The bacterial community structure, in situ spatial distributions and activities of nitrifying and denitrifying bacteria in biofilms treating industrial waste-water were investigated by combination of the 16S rRNA gene clone analysis, fluorescence in situ hybridization (FISH) and microelectrodes. These results were compared with the nitrogen removal capacity of the industrial wastewater treatment plant (IWTP). Both nitrification and denitrification occurred in the primary denitrification (PD) tank and denitrification occurred in the secondary denitrification (SD) tank. In contrast, nitrification and denitrification rates were very low in the nitrification (N) tank. 16S rRNA gene clone sequence analysis revealed that the bacteria affiliated with Alphaproteobacteria, followed by Betaproteobacteria, were numerically important microbial groups in three tanks. The many clones affiliated with Alphaproteobacteria were closely related to the denitrifying bacteria (e.g., Hyphomicrobium spp., Rhodopseudomonas palustris, and Rhodobacter spp.). In addition, Methylophilus leisingeri affiliated with Betaproteobacteria, which favorably utilized methanol, was detected only in the SD-tank to which methanol was added. Nitrosomonas europaea and Nitrosomonas marina were detected as the ammonia-oxidizing bacteria affiliated with Betaproteobacteria throughout this plant, although the dominant species of them was different among three tanks. Nitrifying bacteria were mainly detected in the upper parts of the PD-biofilm whereas their populations were low in the upper parts of the N-biofilm. The presence of denitrifying bacteria affiliated with Hyphomicrobium spp. in SD- and N-biofilms was verified by FISH analysis. Microelectrode measurements showed that the nitrifying bacteria present in the N- and PD-biofilms were active and the bacteria present in the SD-biofilm could denitrify. (c) 2006 Wiley Periodicals, Inc.
  • Olga Savichtcheva, Satoshi Okabe WATER RESEARCH 40 (13) 2463 -2476 2006年07月 [査読無し][通常論文]
     
    The ecological and survival characteristics of bacterial, viral and parasitic pathogens vary under environmental conditions, indicating that probably no single indicator organism can predict the presence of all enteric pathogens for all types of waters and different host-associated fecal pollution. if there are true correlations between indicator organisms and pathogens, it is necessary to find out to what extent and under which circumstances these organisms can be used as reliable indicators of fecal pollution. Application of conventional and alternative fecal indicators has greatly enhanced our abilities to predict and reduce health risk associated with the use of surface waters. New molecular-based techniques have shown that combined use of conventional and alternative indicators for fecal pollution increases both the detection sensitivity and specificity of fecal pollution and associated pathogens. in this review, we, therefore, summarize the advantages and limitations of conventional and alternative fecal indicators in terms of predicting pathogen presence as well as current and future methodologies for direct pathogen monitoring in environmental waters. This manuscript is mainly focused on the relationships between microbial fecal indicators and the presence of pathogens, which have not previously been summarized yet and could nicely supplement with recent literature reviews on microbial source tracking. (c) 2006 Elsevier Ltd. All rights reserved.
  • Y Nakamura, H Satoh, T Kindaichi, S Okabe ENVIRONMENTAL SCIENCE & TECHNOLOGY 40 (5) 1532 -1539 2006年03月 [査読無し][通常論文]
     
    The community structure, spatial distributions, and in situ activity of ammonia-oxidizing bacteria (AOB) representing the Betaproteobacteria and nitrite-oxidizing bacteria (NOB) representing the genus Nitrospira in three different river sediments with different pollution sources and levels along the Niida River, Hachinohe, Japan, were investigated by the combined use of 16S rRNA gene-cloning analysis, real-time quantitative polymerase chain reaction (RTQ-PCR) assays, and microelectrodes. The goal of this research was to evaluate the contribution of nitrifying activity in the sediment to the overall nitrogen elimination rate in this river. The 16S rRNA gene-cloning analysis revealed that the community structures of AOB and Nitrospira-like NOB are present in three sediments. On the basis of the results of 16S rRNA gene-cloning analysis, the RTQ-PCR assay using a TaqMan probe was developed and optimized for the quantification of the Nitrospira-like NOB. In the sediments, AOB specific 16S rRNA genes were detected in the range of 10(6) to 10(7) copies/cm(3) and evenly distributed over the sampled sediment depth (0-5 mm), whereas the Nitrospira-like NOB 16S rRNA gene copy numbers per cm(3) were 1-2 orders of magnitude higher than the AOB copy numbers. Under light conditions, intensive oxygenic photosynthesis occurred in the surface and increased the maximal O-2 concentration and O-2 penetration depth in all sediments. This concomitantly stimulated nitrifying bacteria present in diurnally anoxic deeper zones and expanded nitrification zones, which consequently increased the total NH4+ consumption rate in the sediment (i.e., total NH4+ flux into the sediment). The results suggested that the in situ nitrifying activity was restricted mainly to the surface 2 mm of the sediment and linked with photosynthetic activity, which obviously plays an important role in nitrogen elimination in this river.
  • BE Rittmann, M Hausner, F Loffler, NG Love, G Muyzer, S Okabe, DB Oerther, J Peccia, L Raskin, M Wagner ENVIRONMENTAL SCIENCE & TECHNOLOGY 40 (4) 1096 -1103 2006年02月 [査読無し][通常論文]
  • 金田一 智規, 尾崎 則篤, 對馬 育夫, 小笠原 雄二, 岡部 聡 衛生工学シンポジウム論文集 (13) 195 -198 2005年11月16日 [査読無し][通常論文]
     
    第13回衛生工学シンポジウム(平成17年11月17日(木)-18日(金) 北海道大学クラーク会館) . 一般セッション . 6 水処理 . 6-4
  • O Savichtcheva, N Okayama, T Ito, S Okabe BIOTECHNOLOGY AND BIOENGINEERING 92 (3) 356 -363 2005年11月 [査読無し][通常論文]
     
    To evaluate the viability and survival ability of fecal Bacteroides spp. in environmental waters, a fluorescence-based live/dead staining method using ViaGram (TM) Red(+) Bacterial gram stain and viability kit was combined with fluorescent in situ hybridization (FISH) with 16S rRNA-targeted oligonucleotide probe (referred as ILDS-FISH). The proposed LDS-FISH was a direct and reliable method to detect fecal Bacteroides cells and their viability at single-cell level in complex microbial communities. The pure culture of Bacteroides fragilis and whole human feces were dispersed in aerobic drinking water and incubated at different water temperatures (4 degrees C, 13 degrees C, 18 degrees C, and 24 degrees C), and then the viability of B. fragilis and fecal Bacteroides spp. were determined by applying the LDS-FISH. The results revealed that temperature and the presence of oxygen have significant effects on the survival ability. Increasing the temperature resulted in a rapid decrease in the viability of both pure cultured B. fragilis cells and fecal Bacteroides spp. The live pure cultured B. fragilis cells could be found at the level of detection in drinking water for 48 h of incubation at 24 degrees C, whereas live fecal Bacteroides spp. could be detected for only 4 h of incubation at 24 degrees C. The proposed LDS-FISH method should provide useful quantitative information on the presence and viability of Bacteroides spp., a potential alternative fecal indicator, in environmental waters. (c) 2005 Wiley Periodicals, Inc.
  • A Jang, S Okabe, Y Watanabe, IS Kim, PL Bishop JOURNAL OF ENVIRONMENTAL ENGINEERING AND SCIENCE 4 (5) 413 -420 2005年09月 [査読無し][通常論文]
     
    Optimization of the nitrification processes in biofilms is important for effective nitrogen removal because nitrification in an aerobic biofilm is considered to be a less than reliable process. Thus, one of the main factors to improve biological nitrogen removal processes is a better understanding of the microbiology and population dynamics of ammonia oxidizing bacteria (AOB) in wastewater treatment biofilms. Although the AOB in wastewater treatment have been qualitatively and quantitatively studied, information on their actual populations and activities is still limited. Therefore, the areal cell density of AOB in domestic wastewater biofilms on a partially submerged rotating biological contactor (RBC) was determined by fluorescent in situ hybridization (FISH) with a set of 16S rRNA-targeted oligonucleotide probes. The growth kinetics of the in situ AOB was also studied. Although low numbers of AOB were found at the deeper layers where oxygen was depleted, they were primarily detected in the upper and middle layers of the biofilm. The maximum specific growth rate (mu(b,max)) and half saturation constant (K-s) of AOB in the biofilm were 0.32 d(-1) and 1.7 mM/L of NH4+, respectively.
  • Satoshi Okabe, Tomonori Kindaichi, Tsukasa Ito Applied and environmental microbiology 71 (7) 3987 -94 2005年07月 [査読有り][通常論文]
     
    The cross-feeding of microbial products derived from 14C-labeled nitrifying bacteria to heterotrophic bacteria coexisting in an autotrophic nitrifying biofilm was quantitatively analyzed by using microautoradiography combined with fluorescence in situ hybridization (MAR-FISH). After only nitrifying bacteria were labeled with [14C]bicarbonate, biofilm samples were incubated with and without NH4+ as a sole energy source for 10 days. The transfer of 14C originally incorporated into nitrifying bacterial cells to heterotrophic bacteria was monitored with time by using MAR-FISH. The MAR-FISH analysis revealed that most phylogenetic groups of heterotrophic bacteria except the beta-Proteobacteria showed significant uptake of 14C-labeled microbial products. In particular, the members of the Chloroflexi were strongly MAR positive in the culture without NH4+ addition, in which nitrifying bacteria tended to decay. This indicated that the members of the Chloroflexi preferentially utilized microbial products derived from mainly biomass decay. On the other hand, the members of the Cytophaga-Flavobacterium cluster gradually utilized 14C-labeled products in the culture with NH4+ addition in which nitrifying bacteria grew. This result suggested that these bacteria preferentially utilized substrate utilization-associated products of nitrifying bacteria and/or secondary metabolites of 14C-labeled structural cell components. Our results clearly demonstrated that the coexisting heterotrophic bacteria efficiently degraded and utilized dead biomass and metabolites of nitrifying bacteria, which consequently prevented accumulation of organic waste products in the biofilm.
  • T Ito, K Sugita, Yumoto, I, Y Nodasaka, S Okabe INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY 55 (3) 1059 -1064 2005年05月 [査読無し][通常論文]
     
    A novel mesophilic, chemolithoautotrophic, sulfur-oxidizing bacterium, designated strain SO07(T) was isolated from a microaerobic waste-water biofilm. Chemolithoautotrophic growth was observed with elemental sulfur, sulfide and thiosulfate as sole electron donors and oxygen as electron acceptor. Anaerobic and heterotrophic growth were not observed. Nitrate was not used as a terminal electron acceptor. The optimum pH and temperature for growth were pH 7(.)5 and 30 degrees C, respectively. The major isoprenoid quinone was Q-8. The DNA G + C content of strain SO07(T) was 47(.)1 mol%. Phylogenetic analysis of 16S rRNA gene sequences demonstrated that strain SO07(T) formed a monophyletic group in the gamma-Proteobacteria with only 89% similarity to members of the genus Halothiobacillus, its nearest phylogenetic neighbours. In addition, the isolate differed from members of the genus Halothiobacillus in its requirement for and tolerance of NaCl; strain SO07(T) was unable to grow in NaCl concentrations of more than 180 mM. On the basis of phylogenetic, chemotaxonomic and physiological data, it is proposed that isolate SO07(T) (= JCM 12417(T) = ATCC BAA- 1033(T)) represents the type strain of a novel species in a new genus, Thiovirga sulfuroxydans gen. nov., sp. nov.
  • Satoshi Okabe, Tsukasa Ito, Kenichi Sugita, Hisashi Satoh Applied and Environmental Microbiology 71 (5) 2520 -2529 2005年05月 [査読無し][通常論文]
     
    The succession of sulfur-oxidizing bacterial (SOB) community structure and the complex internal sulfur cycle occurring in wastewater biofilms growing under microaerophilic conditions was analyzed by using a polyphasic approach that employed 16S rRNA gene-cloning analysis combined with fluorescence in situ hybridization, microelectrode measurements, and standard batch and reactor experiments. A complete sulfur cycle was established via S0 accumulation within 80 days in the biofilms in replicate. This development was generally split into two phases, (i) a sulfur-accumulating phase and (ii) a sulfate-producing phase. In the first phase (until about 40 days), since the sulfide production rate (sulfate-reducing activity) exceeded the maximum sulfide-oxidizing capacity of SOB in the biofilms, H2S was only partially oxidized to S0 by mainly Thiomicrospira denitirificans with NO3- as an electron acceptor, leading to significant accumulation of S0 in the biofilms. In the second phase, the SOB populations developed further and diversified with time. In particular, S 0 accumulation promoted the growth of a novel strain, strain SO07, which predominantly carried out the oxidation of S0t to SO 42- under oxic conditions, and Thiothrix sp. strain CT3. In situ hybridization analysis revealed that the dense populations of Thiothrix (ca. 109 cells cm-3) and strain SO07 (ca. 108 cells cm-3) were found at the sulfur-rich surface (100 μm), while the population of Thiomicrospira denitirificans was distributed throughout the biofilms with a density of ca. 107 to 108 cells cm -3. Microelectrode measurements revealed that active sulfide-oxidizing zones overlapped the spatial distributions of different phylogenetic SOB groups in the biofilms. As a consequence, the sulfide-oxidizing capacities of the biofilms became high enough to completely oxidize all H 2S produced by SRB to SO42- in the second phase, indicating establishment of the complete sulfur cycle in the biofilms. Copyright © 2005, American Society for Microbiology. All Rights Reserved.
  • 三浦佑己, 伊藤司, 渡辺義公, 岡部聡 日本水環境学会年会講演集 39th 41 2005年03月17日 [査読無し][通常論文]
  • 中村吉志, 渡辺義公, 岡部聡, 佐藤久 日本水環境学会年会講演集 39th 231 2005年03月17日 [査読無し][通常論文]
  • 對馬 育夫, 小笠原 雄二, 金田一 智規, 岡部 聡 日本微生物生態学会講演要旨集 (21) 206 -206 2005年 [査読無し][通常論文]
  • S Okabe, T Kindaichi, Y Nakamura, T Ito WATER SCIENCE AND TECHNOLOGY 52 (7) 225 -232 2005年 [査読無し][通常論文]
     
    Microautoradiography combined with fluorescent in situ hybridization (MAR-FISH), a powerful tool for linking physiology with identification of individual cells, was applied to investigate microbial interactions between nitrifying bacteria and coexisting heterotrophic bacteria in an autotrophic nitrifying biofilm community fed with only ammonia as the sole energy source and bicarbonate as the sole carbon source. First, nitrifying bacteria were radiolabeled by culturing the biofilm samples with [C-14]bicarbonate for 6 h, and then the transfer of radioactivity from nitrifying bacteria to heterotrophic bacteria was monitored by using MAR-FISH. MAR-FISH revealed that the heterotrophic bacterial community was composed of bacteria that were phylogenetically and metabofically diverse. We could obtain direct evidence that organic matter derived from nitrifiers was subsequently utilized by mainly filamentous bacteria belonging to the Chloroflexi (green non-sulfur bacteria) group or CFB group in the biofilm, which was clearly visualized by MAR-FISH at single cell resolution for the first time. On the other hand, the members of the alpha- and gamma-Proteobacteria were specialized to utilize low-molecular-weight organic matter. This community represents functionally integrated units that assure maximum access to and utilization of metabolites of nitrifiers.
  • 微生物群集構造解析技術の現状と今後の展開・課題
    水環境学会誌 28 (8) 460 -465 2005年 [査読無し][通常論文]
  • T Ito, K Sugita, S Okabe APPLIED AND ENVIRONMENTAL MICROBIOLOGY 70 (5) 3122 -3129 2004年05月 [査読無し][通常論文]
     
    We successfully isolated a novel aerobic chemolithotrophic sulfur-oxidizing bacterium, designated strain SO07, from wastewater biofilms growing under microaerophilic conditions. For isolation, the use of elemental sulfur (S-0), which is the most abundant sulfur pool in the wastewater biofilms, as the electron donor was an effective measure to establish an enrichment culture of strain SO07 and further isolation. 16S rRNA gene sequence analysis revealed that newly isolated strain SO07 was affiliated with members of the genus Halothiobacillus, but it was only distantly related to previously isolated species (89% identity). Strain SO07 oxidized elemental sulfur, thiosulfate, and sulfide to sulfate under oxic conditions. Strain SO07 could not grow on nitrate. Organic carbons, including acetate, propionate, and formate, could not serve as carbon and energy sources. Unlike other aerobic sulfur-oxidizing bacteria, this bacterium was sensitive to NaCl; growth in medium containing more than 150 mM was negligible. In situ hybridization combined with confocal laser scanning microscopy revealed that a number of rod-shaped cells hybridized with a probe specific for strain SO07 were mainly present in the oxic biofilm strata (ca. 0 to 100 mum) and that they often coexisted with sulfate-reducing bacteria in this zone. These results demonstrated that strain SO07 was one of the important sulfur-oxidizing populations involved in the sulfur cycle occurring in the wastewater biofilm and was primarily responsible for the oxidation of H2S and S-0 to SO42- under oxic conditions.
  • 巽善彦, 渡辺義公, 岡部聡 日本水環境学会年会講演集 38th 196 2004年03月17日 [査読無し][通常論文]
  • 久保広明, 渡辺義公, 岡部聡, 木村克輝 日本水環境学会年会講演集 38th 363 2004年03月17日 [査読無し][通常論文]
  • 岡山紀子, SAVICHTCHEVA O, 金田一智規, 伊藤司, 岡部聡, 渡辺義公 日本水環境学会年会講演集 38th 313 2004年03月17日 [査読無し][通常論文]
  • 金田一智規, 伊藤司, 岡部聡, 渡辺義公 日本水環境学会年会講演集 38th 380 2004年03月17日 [査読無し][通常論文]
  • 対馬育夫, 金田一智規, 岡部聡, 渡辺義公 日本水環境学会年会講演集 38th 370 2004年03月17日 [査読無し][通常論文]
  • 中村吉志, 岡部聡, 渡辺義公, 佐藤久 日本水環境学会年会講演集 38th 508 2004年03月17日 [査読無し][通常論文]
  • H Satoh, H Ono, B Rulin, J Kamo, S Okabe, KI Fukushi WATER RESEARCH 38 (6) 1633 -1641 2004年03月 [査読無し][通常論文]
     
    A membrane aerated biofilm reactor (MABR), in which O-2 was supplied from the bottom of the biofilm and NH4+ and organic carbon were supplied from the biofilm surface, was operated at different organic carbon loading rates and intra-membrane air pressures to investigate the occurrence of simultaneous chemical oxygen demand (COD) removal, nitrification and denitrification. The spatial distribution of nitrification and denitrification zones in the biofilms was measured with microelectrodes for O-2, NH4+, NO2-, NO3- and pH. When the MABR was operated at approximately 1.0 g-COD/m(2)/day of COD loading rate, simultaneous COD removal, nitrification and denitrification could be achieved. The COD loading rates and the intra-membrane air pressures applied in this study had no effect on the startup and the maximum rates of NH4+ oxidation in the MABRs. Microelectrode measurements showed that O-2 was supplied from the bottom of the MABR biofilm and penetrated the whole biofilm. Because the biofilm thickness increased during the operations, an anoxic layer developed in the upper parts of the mature biofilms while an oxic layer was restricted to the deeper parts of the biofilms. The development of the anoxic zones in the biofilms coincided with increase in the denitrification rates. Nitrification occurred in the zones from membrane surface to a point of ca. 60 mum. Denitrification mainly occurred just above the nitrification zones. The COD loading rates and the intra-membrane air pressures applied in this study had no effect on location of the nitrification and denitrification zones. (C) 2004 Elsevier Ltd. All rights reserved.
  • Satoshi Okabe, Tomonori Kindaichi, Tsukasa Ito, Hisashi Satoh Biotechnology and Bioengineering 85 (1) 86 -95 2004年01月05日 [査読無し][通常論文]
     
    A fine-scale in situ spatial organization of ammonia-oxidizing bacteria (AOB) in biofilms was investigated by combining molecular techniques (i.e., fluorescence in situ hybridization (FISH) and 16S rDNA-cloning analysis) and microelectrode measurements. Important parameters of AOB microcolonies such as size distribution and areal cell density of the microcolonies were determined and correlated with substrate microprofiles in the biofilms. In situ hybridization with a nested 16S rRNA-targeted oligonucleotide probe set revealed two different populations of AOB, Nitrosomonas europaea-lineage and Nitrosospira multiformis-lineage, coexisting in an autotrophic nitrifying biofilm. Nitrosospira formed looser microcolonies, with an areal cell density of 0.51 cells μm-2, which was half of the cell density of Nitrosomonas (1.12 cells μm-2). It is speculated that the formation of looser microcolonies facilitates substrate diffusion into the microcolonies, which might be a survival strategy to low O2 and NH4+ conditions in the biofilm. A long-term experiment (4-week cultivation at different substrate C/N ratios) revealed that the size distribution of AOB microcolonies was strongly affected by better substrate supply due to shorter distance from the surface and the presence of organic carbon. The microcolony size was relatively constant throughout the autotrophic nitrifying biofilm, while the size increased by ∼ 80% toward the depth of the biofilm cultured at the substrate C/N = 1. A short-term (∼ 3 h) organic carbon addition experiment showed that the addition of organic carbon created interspecies competition for O2 between AOB and heterotrophic bacteria, which dramatically decreased the in situ NH4 +-uptake activity of AOB in the surface of the biofilms. This result might explain the spatial distribution of AOB microcolony size in the biofilms cultured at the substrate C/N = 1. These experimental results suggest O 2 and organic carbon were the main factors controlling the spatial organization and activity of AOB in biofilms. These findings are significantly important to further improve mathematical models used to describe how the slow-growing AOB develop their niches in biofilms and how that configuration affects nitrification performance in the biofilm. © 2004 Wiley Periodicals, Inc.
  • T Kindaichi, S Okabe, H Satoh, Y Watanabe WATER SCIENCE AND TECHNOLOGY 49 (11-12) 61 -68 2004年 [査読無し][通常論文]
     
    Effects of hydroxylamine (NH2OH), an intermediate of NH4+ oxidation, on microbial community structure and function of two autotrophic nitrifying biofilms fed with and without NH2OH were analyzed by a 16S rRNA approach and the use of microelectrodes. In the NH2OH-added biofilm, partial oxidation of NH4+ to NO2- was observed, whereas complete oxidation of NH4+ to NO3- was achieved in the control biofilm. In situ hybridization results revealed that no nitrite-oxidizing bacteria (NOB) hybridized with any specific probes were detected in the NH2OH-added biofilm. Thus, the addition of low concentrations of NH2OH (250 muM) completely inhibited the growth of NOR Phylogenetic analysis of 16S rDNA indicated that the ammonia-oxidizing bacteria (AOB) detected in both biofilms were closely related to Nitrosomonas europaea, and that the clone sequences from both biofilm libraries have more than 99% similarity to each other. However, in situ hybridization results revealed that the addition of NH2OH changed the form of growth pattern of the dominant Nitrosomonas spp. from dense clusters mode to single scattered cells mode. Microelectrode measurements revealed that the average NH4+ consumption rate calculated in the NH2OH-added biofilm was two times higher than that in the control biofilm. This clearly demonstrated that the oxidation of NH4+ was stimulated by NH2OH addition.
  • 渡辺義公, 岡部聡, 木村克輝 第4回廃棄物対策研究発表会成果発表抄録集 平成16年度廃棄物対策研究推進事業 I.35-I.37 2004年 [査読無し][通常論文]
  • T. Kindaichi, S. Okabe, H. Satoh, Y. Watanabe Water Science and Technology 49 (11-12) 61 -68 2004年 [査読無し][通常論文]
     
    Effects of hydroxylamine (NH2OH), an intermediate of NH4+ oxidation, on microbial community structure and function of two autotrophic nitrifying biofilms fed with and without NH2OH were analyzed by a 16S rRNA approach and the use of microelectrodes. In the NH2OH-added biofilm, partial oxidation of NH4+ to NO2- was observed, whereas complete oxidation of NH4+ to NO3- was achieved in the control biofilm. In situ hybridization results revealed that no nitrite-oxidizing bacteria (NOB) hybridized with any specific probes were detected in the NH2OH-added biofilm. Thus, the addition of low concentrations of NH2OH (250 μM) completely inhibited the growth of NOB. Phylogenetic analysis of 16S rDNA indicated that the ammonia-oxidizing bacteria (AOB) detected in both biofilms were closely related to Nitrosomonas europaea, and that the clone sequences from both biofilm libraries have more than 99% similarity to each other. However, in situ hybridization results revealed that the addition of NH2OH changed the form of growth pattern of the dominant Nitrosomonas spp. from dense clusters mode to single scattered cells mode. Microelectrode measurements revealed that the average NH4+ consumption rate calculated in the NH2OH-added biofilm was two times higher than that in the control biofilm. This clearly demonstrated that the oxidation of NH4+ was stimulated by NH2OH addition. © IWA Publishing 2004.
  • S Wuertz, S Okabe, M Hausner WATER SCIENCE AND TECHNOLOGY 49 (11-12) 327 -336 2004年 [査読無し][通常論文]
     
    Several important advances have been made in the study of biofilm microbial populations relating to their spatial structure (or architecture), their community structure, and their dependence on physicochemical parameters. With the knowledge that hydrodynamic forces influence biofilm architecture came the realization that metabolic processes may be enhanced if certain spatial structures can be forced. An example is the extent of plasmid-mediated horizontal gene transfer in biofilms. Recent in situ work in defined model systems has shown that the biofilm architecture plays a role for genetic transfer by bacterial conjugation in determining how far the donor cells can penetrate the biofilm. Open channels and pores allow for more efficient donor transport and hence more frequent cell collisions leading to rapid spread of the genes by horizontal gene transfer. Such insight into the physical environment of biofilms can be utilized for bioenhancement of catabolic processes by introduction of mobile genetic elements into an existing microbial community. If the donor organisms themselves persist, bioaugmentation can lead to successful establishment of newly introduced species and may be a more successful strategy than biostimulation (the addition of nutrients or specific carbon sources to stimulate the authochthonous population) as shown for an enrichment culture of nitrifying bacteria added to rotating disk biofilm reactors using fluorescent in situ hybridization (FISH) and microelectrode measurements of NH4+, NO2-, NO3-, and O-2. However, few studies have been carried out on full-scale systems. Bioaugmentation and bioenhancement are most successful if a constant selective pressure can be maintained favoring the promulgation of the added enrichment culture. Overall, knowledge gain about microbial community interactions in biofilms continues to be driven by the availability of methods for the rapid analysis of microbial communities and their activities. Molecular tools can be grouped into those suitable for ex situ and in situ community analysis. Non-spatial community analysis, in the sense of assessing changes in microbial populations as a function of time or environmental conditions, relies on general fingerprinting methods, like DGGE and T-RFLP, performed on nucleic acids extracted from biofilm. These approaches have been most useful when combined with gene amplification, cloning and sequencing to assemble a phylogenetic inventory of microbial species. It is expected that the use of oligonucleotide microarrays will greatly facilitate the analysis of microbial communities and their activities in biofilms. Structure-activity relationships can be explored using incorporation of C-13-labeled substrates into microbial DNA and RNA to identify metabolically active community members. Finally, based on the DNA sequences in a biofilm, FISH probes can be designed to verify the abundance and spatial location of microbial community members. This in turn allows for in situ structure/function analysis when FISH is combined with microsensors, microautoradiography, and confocal laser scanning microscopy with advanced image analysis.
  • Photosynthesis in sediment at a tidal area of Niida River, Hachinohe, Japan
    Water Research 38 (9) 2439 -2447 2004年 [査読無し][通常論文]
  • KINDAICHI T, ITO T, OKABE S Applied and Environmental Microbiology 70 (3) 1641 -1650 2004年 [査読無し][通常論文]
  • 金田一 智規, 河野 快子, 伊藤 司, 岡部 聡 環境工学研究論文集 41 (41) 321 -330 2004年 [査読無し][通常論文]
     
    We investigated the population dynamics of nitrifying bacteria in autotrophic nitrifying biofilms by using Real-Time PCR and fluorescent in situ hybridization (FISH). Primers and TaqMan probe specific to genus Nitrospira including uncultured Nitrospira sp. in the biofilms were newly designed and applied to the biofilms. The populations of ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB) determined by Real-Time PCR during steady-state condition were 1.2×108cells/cm2 and 3.5×108cells/cm2, respectively. The population dynamics of nitrifying bacteria corresponded with the reactor performance and the development of nitrifying bacteria in the biofilms determined by FISH. Real-Time PCR revealed that specific growth rates of uncultured Nitrospira sp. were 0.021h-1 in the suspended enrichment culture and 0.014h-1 in the biofilms. Average specific growth rates of both nitrifying bacteria during the steady-state condition in the biofilms were significantly lower than that during the log phase, might suggest that metabolic pathway of nitrifying bacteria in the biofilms was different in the log and stationary phase. We speculated that Real-Time PCR was powerful and suitable technique for understanding of microbial population dynamics in the complex microbial communities.
  • バイオフィルム内の細菌の特異的検出とその分布の測定
    Bacterial Adherence & Biofilm 17 102 -107 2004年 [査読無し][通常論文]
  • S Okabe, T Ito, H Satoh APPLIED MICROBIOLOGY AND BIOTECHNOLOGY 63 (3) 322 -334 2003年12月 [査読無し][通常論文]
     
    The community structure of sulfate-reducing bacteria (SRB) and the contribution of SRB to carbon mineralization in a wastewater biofilm growing under microaerophilic conditions were investigated by combining molecular techniques, molybdate inhibition batch experiments, and microelectrode measurements. A 16S rDNA clone library of bacteria populations was constructed from the biofilm sample. The 102 clones analyzed were grouped into 53 operational taxonomic units (OTUs), where the clone distribution was as follows: Cytophaga-Flexibacter-Bacteroides (41%), Proteobacteria (41%), low-G+C Gram-positive bacteria (18%), and other phyla (3%). Three additional bacterial clone libraries were also constructed from SRB enrichment cultures with propionate, acetate, and H-2 as electron donors to further investigate the differences in SRB community structure due to amendments of different carbon sources. These libraries revealed that SRB clones were phylogenetically diverse and affiliated with six major SRB genera in the delta-subclass of the Proteobacteria. Fluorescent in situ hybridization (FISH) analysis revealed that Desulfobulbus and Desulfonema were the most abundant SRB species in this biofilm, and this higher abundance (ca. 2-4x10(9) cells cm(-3) and 5x10(7) filaments cm(-3), respectively) was detected in the surface of the biofilm. Microelectrode measurements showed that a high sulfate-reducing activity was localized in a narrow zone located just below the oxic/anoxic interface when the biofilm was cultured in a synthetic medium with acetate as the sole carbon source. In contrast, a broad sulfate-reducing zone was found in the entire anoxic strata when the biofilm was cultured in the supernatant of the primary settling tank effluent. This is probably because organic carbon sources diffused into the biofilm from the bulk water and an unknown amount of volatile fatty acids was produced in the biofilm. A combined approach of molecular techniques and batch experiments with a specific inhibitor (molybdate) clearly demonstrated that Desulfobulbus is a numerically important member of SRB populations and the main contributor to the oxidation of propionate to acetate in this biofilm. However, acetate was preferentially utilized by nitrate-reducing bacteria but not by acetate-utilizing SRB.
  • 久保広明, 岡部聡, 木村克輝, 渡辺義公 土木学会年次学術講演会講演概要集(CD-ROM) 58th (Disk 2) VII-085 2003年09月01日 [査読無し][通常論文]
  • 岡部聡, 小田切光典, 伊藤司, 佐藤久, 小山田浩之, 渡辺義公 土木学会年次学術講演会講演概要集(CD-ROM) 58th (Disk 2) VII-189 2003年09月01日 [査読無し][通常論文]
  • H Satoh, Y Nakamura, H Ono, S Okabe BIOTECHNOLOGY AND BIOENGINEERING 83 (5) 604 -607 2003年09月 [査読無し][通常論文]
     
    Simultaneous nitrification and denitrification (SND) was investigated in the single aeration tank of a municipal wastewater treatment plant. Microelectrode measurements and batch experiments were performed to test for the presence of SND. Microelectrodes recorded the presence of O-2 concentration gradients in individual activated sludge flocs. When the O-2 concentration in the bulk liquid was <45 μM, anoxic zones were detected within flocs with a larger diameter (approximately 3000 pm). The O-2 penetration depth in the floc was found to be dependent on the O-2 concentration in the bulk liquid. Nitrification was restricted to the oxic zones, whereas denitrification occurred mainly in the anoxic zones. The nitrification rate of the activated sludge increased with increasing O-2 concentration in the bulk liquid, up to 40 μM, and remained constant thereafter. SND was observed in the aerated activated sludge when O-2 concentration was in the range of 10 to 35 μM. (C) 2003 Wiley Periodicals, Inc.
  • GH Chen, MT Wong, S Okabe, Y Watanabe WATER RESEARCH 37 (13) 3125 -3135 2003年07月 [査読無し][通常論文]
     
    Dynamic response of nitrifying activated sludge batch cultures to increased chloride concentration was studied in this paper, which focused upon the changes in the specific nitrification rate (SNR) and nitrifier population when the chloride level was gradually or stepwise increased to 30,000 mg Cl L-1. The dominant species of ammonia-oxidizers and nitrite-oxidizers in the population were examined by Fluorescent in situ hybridization technique with 16S rRNA-targeted oligonucleotide probes. It was found that neither chloride increasing approaches affected the SNR of the batch cultures before the chloride concentration exceeded 10,000 mg Cl L-1, after which the stepwise increase approach reduced the SNR more significantly than the gradual increase approach. From 10,000 to 18,000 mg Cl L-1 a down-and-up pattern of the SNR variation appeared in both approaches, which was associated with the change in the dominant species of ammonia-oxidizers from non-saline-resistant species such as Nitrosomonas europaea-lineage and Nitrosomonas eutropha to saline-resistant species, such as the Nitrosococcus mobilis-lineage. Nitrobacter was the only dominant species when the chloride concentration was below 10,000 mg Cl L-1, where no nitrite-oxidizers survived. Therefore, the 10,000 mg Cl L-1 chloride level is a critical level for the shift of the nitrifier population in the nitrifying activated sludge batch cultures. (C) 2003 Elsevier Science Ltd. All rights reserved.
  • H Satoh, S Okabe, Y Yamaguchi, Y Watanabe WATER RESEARCH 37 (9) 2206 -2216 2003年05月 [査読無し][通常論文]
     
    Three rotating disk biofilm reactors were operated to evaluate whether bioaugmentation and biostimulation can be used to improve the start-up of microbial nitrification. The first reactor was bioaugmented during start-up period with an enrichment culture of nitrifying bacteria, the second reactor received a synthetic medium containing NH4+ and NO2- to facilitate concomitant proliferation of ammonia- and nitrite-oxidizing bacteria, and the third reactor was used as a control. To evaluate the effectiveness of bioaugmentation and biostimulation approaches, time-dependent developments of nitrifying bacterial community and in situ nitrifying activity in biofilms were monitored by fluorescence in situ hybridization (FISH) technique and microelectrode measurements of NH4+, NO2-, NO3-, and O-2. In situ hybridization results revealed that addition of the enrichment culture of nitrifying bacteria significantly facilitated development of dense nitrifying bacterial populations in the biofilm shortly after, which led to a rapid start-up and enhancement of in situ nitrification activity. The inoculated bacteria could proliferate and/or survive in the biofilm. In addition, the addition of nitrifying bacteria increased the abundance of nitrifying bacteria in the surface of the biofilm, resulting in the higher nitrification rate. On the other hand, the addition of 2.1 mM NO2- did not stimulate the growth of nitrite-oxidizing bacteria and did inhibit the proliferation of ammonia-oxidizing bacteria instead. Thus, the start-up of NO2- oxidation was unchanged, and the start-up of NH4+ oxidation was delayed. In all the three biofilm reactors, data sets of time series analyses on population dynamics of nitrifying bacteria determined by FISH, in situ nitrifying activities determined by microelectrode measurements, and the reactor performances revealed an approximate agreement between the appearance of nitrifying bacteria and the initiation of nitrification activity, suggesting that the combination of these techniques was a very powerful monitoring tool to evaluate the effectiveness of bioaugmentation and biostimulation strategies. (C) 2002 Elsevier Science Ltd. All rights reserved.
  • 金田一智規, 伊藤司, 岡部聡, 渡辺義公 日本水環境学会年会講演集 37th 62 2003年03月04日 [査読無し][通常論文]
  • 対島育夫, 金田一智規, 岡部聡, 渡辺義公 日本水環境学会年会講演集 37th 507 2003年03月04日 [査読無し][通常論文]
  • 杉田謙一, 伊藤司, 岡部聡, 渡辺義公 日本水環境学会年会講演集 37th 67 2003年03月04日 [査読無し][通常論文]
  • AM Briones, S Okabe, Y Umemiya, NB Ramsing, W Reichardt, H Okuyama PLANT AND SOIL 250 (2) 335 -348 2003年03月 [査読無し][通常論文]
     
    Ammonia-oxidizing bacteria (AOB) populations were studied on the root surface of different rice cultivars by PCR coupled with denaturing gradient gel electrophoresis (DGGE) and fluorescence in situ hybridization (FISH). PCR-DGGE of the ammonium monooxygenase gene (amoA) showed a generally greater diversity on root samples compared to rhizosphere and unplanted soil. Sequences affiliated with Nitrosomonas spp. tended to be associated with modern rice hybrid lines. Root-associated AOB observed by FISH were found within a discrete biofilm coating the root surface. Although the total abundance of AOB on root biofilms of different rice cultivars did not differ significantly, there were marked contrasts in their population structure, indicating selection of Nitrosomonas spp. on roots of a hybrid cultivar. Observations by FISH on the total bacterial community also suggested that different rice cultivars support different bacterial populations even under identical environmental conditions. The presence of active AOB in the root environment predicts that a significant proportion of the N taken up by certain rice cultivars is in the form of NO3--N produced by the AOB. Measurement of plant growth of hydroponically grown plants showed a stronger response of hybrid cultivars to the co-provision of NH4+ and NO3-. In soil-grown plants, N use efficiency in the hybrid was improved during ammonium fertilization compared to nitrate fertilization. Since ammonium-fertilized plants actually receive a mixture of NH4+ and NO3- with ratios depending on root-associated nitrification activity, these results support the advantage of co-provision of ammonium and nitrate for the hybrid cultivar.
  • S Okabe, CM Santegoeds, D De Beer BIOTECHNOLOGY AND BIOENGINEERING 81 (5) 570 -577 2003年03月 [査読無し][通常論文]
     
    Microelectrode, fluorescence in situ hybridization (FISH), and denaturing gradient gel electrophoresis (DGGE) analyses were used to investigate the effect of nitrite and nitrate on in situ sulfide production in an ac activated sludge immobilized agar gel film. Microelectrode measurements of O-2, H2S, NO3-, NO2-, and pH revealed that the addition of NO2- and NO3- forced sulfate reduction zones deeper in the agar gel and significantly reduced the in situ sulfide production levels. The sulfate reduction zone was consequently separated from 02 and NO2- or NO3- respiration zones with increasing the concentrations of NO2- and NO3-. These NO2- and NO3- treatments had only a transient effect on sulfide production. The in situ sulfide production quickly recovered to the previous levels when NO2- and NO3- were removed. The PCR-DGGE and FISH analyses revealed that 2-day-continuous addition of 500 muM NO3- did not change the metabolically active sulfate-reducing bacterial (SRB) community. On the basis of these data, it could be concluded that the addition of NO2- and NO3- did not kill SRB, but induced the interspecies competition for common carbon source (i.e., acetate) between nitrate-reducing heterotrophic bacteria and SRB and enhanced the oxidation of the produced sulfide, which were main possible causes of the suppression of in situ sulfide production in the agar gel. (C) 2003 Wiley Periodicals, Inc.
  • A Jang, PL Bishop, S Okabe, SG Lee, IS Kim WATER SCIENCE AND TECHNOLOGY 47 (1) 49 -57 2003年 [査読無し][通常論文]
     
    A better understanding of microbiology and ecology of nitrifying bacteria in inner biofilms is an important part of improving process performance and control. Microelectrodes and fluorescent in situ hybridization (FISH) in biofilm research have been used to investigate the spatial distributions of various microbial activities in biofilms and have led to new experimental findings as well as modifications of the homogeneous assumptions in the biofilm kinetic models. The objective of this study is to try the combination of two methods, both FISH and microelectrode measurements, and to provide reliable and in situ information on nitrifying bacterial activity in biofilms. The characteristics of biofilm developed on tygon slides were different according to the change of dissolved oxygen (DO). When the DO increased from 2 to 10 mg DO/L, the rate of the biofilm thickness increased and its dry density changed from 50-70 to 25-90 mg/cm(3). Ammonia oxidizing bacteria were not uniformly distributed in biofilm, and were found at the deeper layer where oxygen is depleted, they were detected primarily in the upper and middle layers of the biofilm.
  • S Okabe, T Ito, H Satoh, Y Watanabe WATER SCIENCE AND TECHNOLOGY 47 (11) 281 -288 2003年 [査読無し][通常論文]
     
    The effects of O-2 and NO3- concentrations on in situ sulfate reduction and sulfide reoxidation in microaerophilic wastewater biofilms grown on rotating disk reactors were investigated by the use of microelectrodes for O-2, S2-, NO3-, NO2-, and pH. Microelectrocle measurements showed the vertical microzonation of O-2 respiration, NO3- respiration, H2S oxidation and SO42- reduction in the biofilms. The microelectrode measurements indicate that sulfate reducing activity was largely restricted to a narrow anaerobic zone located about 500 mum below the biofilm surface. An addition of nitrate forced the sulfate reduction zone deeper in the biofilm and reduced the specific sulfate reduction rate as well. The sulfate reduction zone was consequently separated from the O-2 and NO3- respiration zones. Anaerobic H2S oxidation with NO3- was also induced by addition of nitrate to the medium. Measurements of the reduced inorganic sulfur compounds (FeS, FeS2 and SO), total-Mn and total-Fe in the biofilm indicated that the produced H2S became immediately oxidized with O-2, NO3- and other oxidants, mainly ferric/ferrous hydrates. On the basis of the present results, it was estimated that of all sulfide produced, 13% of the sulfide was precipitated by metal ions as FeS and SO just above the sulfate reduction zone, 65% was anaerobically oxidized to SO42- with NO3- as an electron acceptor and 22% was aerobically oxidized within the biofilm incubated in 70 mumol l(-1) of DO and 280 mumol l(-1) of NO3-.
  • S. Okabe, T. Ito, H. Satoh, Y. Watanabe Water Science and Technology 47 (11) 281 -288 2003年 [査読無し][通常論文]
     
    The effects of O2 and NO3- concentrations on in situ sulfate reduction and sulfide reoxidation in microaerophilic wastewater biofilms grown on rotating disk reactors were investigated by the use of microelectrodes for O2, S2-, NO3-, NO2-, and pH. Microelectrode measurements showed the vertical microzonation of O2 respiration, NO3- respiration, H2S oxidation and SO42- reduction in the biofilms. The microelectrode measurements indicate that sulfate reducing activity was largely restricted to a narrow anaerobic zone located about 500 μm below the biofilm surface. An addition of nitrate forced the sulfate reduction zone deeper in the biofilm and reduced the specific sulfate reduction rate as well. The sulfate reduction zone was consequently separated from the O2 and NO3- respiration zones. Anaerobic H2S oxidation with NO3- was also induced by addition of nitrate to the medium. Measurements of the reduced inorganic sulfur compounds (FeS, FeS2 and S0), total-Mn and total-Fe in the biofilm indicated that the produced H2S became immediately oxidized with O2, NO3- and other oxidants, mainly ferric/ferrous hydrates. On the basis of the present results, it was estimated that of all sulfide produced, 13% of the sulfide was precipitated by metal ions as FeS and S0 just above the sulfate reduction zone, 65% was anaerobically oxidized to SO42- with NO3- as an electron acceptor and 22% was aerobically oxidized within the biofilm incubated in 70 μmol l1- of DO and 280 μmol l1- of NO3-.
  • A Jang, PL Bishop, S Okabe, SG Lee, IS Kim WATER SCIENCE AND TECHNOLOGY 47 (1) 49 -57 2003年 [査読無し][通常論文]
     
    A better understanding of microbiology and ecology of nitrifying bacteria in inner biofilms is an important part of improving process performance and control. Microelectrodes and fluorescent in situ hybridization (FISH) in biofilm research have been used to investigate the spatial distributions of various microbial activities in biofilms and have led to new experimental findings as well as modifications of the homogeneous assumptions in the biofilm kinetic models. The objective of this study is to try the combination of two methods, both FISH and microelectrode measurements, and to provide reliable and in situ information on nitrifying bacterial activity in biofilms. The characteristics of biofilm developed on tygon slides were different according to the change of dissolved oxygen (DO). When the DO increased from 2 to 10 mg DO/L, the rate of the biofilm thickness increased and its dry density changed from 50-70 to 25-90 mg/cm(3). Ammonia oxidizing bacteria were not uniformly distributed in biofilm, and were found at the deeper layer where oxygen is depleted, they were detected primarily in the upper and middle layers of the biofilm.
  • 金田一 智規, 伊藤 司, 岡部 聡, 渡辺 義公 環境工学研究論文集 40 (40) 71 -79 2003年 [査読無し][通常論文]
     
    We investigated the phylogenetic diversity of ammonia-oxidizing bacteria (AOB) in different environments based on 16S ribosomal DNA (rDNA) sequence analysis with the specific primer set. The sequence analysis revealed that the Nitrosomonas oligotropha-like AOB were the dominant in low-ammonium environments such as river and drinking water treatment systems. In contrast, the Nitrosomonas europaea-like AOB were the dominant in high-ammonium environments in which the concentration of ammonium is more than 1000 mg-N/L. In medium-ammonium environments, the coexistence of the distinct AOB related to the Nitrosomonas europaea-, Nitrosomonas oligotropha-, and Nitrosospira sp.-like clusters were found, indicating a high phylogenetic diversity. These observations support the hypothesis that Nitmsomonas europaea is r-strategist, whereas Nitrosospira sp. is K-strategist, indicating that Nitrosomonas europaea can outcompete the Nitrosospira sp. at high ammonium concentration. These results provide insight into the type of AOB responsible for nitrification in different engineered systems, which should help direct future studies aimed at characterizing relevant AOB growth.
  • バイオフィルム内における硫黄循環と関連微生物群集構造の解析
    月刊海洋号外「海洋微生物Ⅱ-基礎,応用研究とその利用」 35 54 -63 2003年 [査読無し][通常論文]
  • 岡部聡, 伊藤司, 佐藤久, 渡辺義公 用水と廃水 44 (11) 961 -970 2002年11月01日 [査読無し][通常論文]
  • 杉田謙一, 岡部聡, 伊藤司, 渡辺義公 土木学会年次学術講演会講演概要集(CD-ROM) 57th VII-284 2002年09月01日 [査読無し][通常論文]
  • AM Briones, S Okabe, Y Umemiya, NB Ramsing, W Reichardt, H Okuyama APPLIED AND ENVIRONMENTAL MICROBIOLOGY 68 (6) 3067 -3075 2002年06月 [査読無し][通常論文]
     
    Comparisons of the activities and diversities of ammonia-oxidizing bacteria (AOB) in the root environment of different cultivars of rice (Oryza sativa L.) indicated marked differences despite identical environmental conditions during growth. Gross nitrification rates obtained by the N-15 dilution technique were significantly higher in a modern variety, IR63087-1-17, than in two traditional varieties. Phylogenetic analysis based on the ammonium monooxygenase gene (amoA) identified strains related to Nitrosospira multiformis and Nitrosomonas europaea as the predominant AOB in our experimental rice system. A method was developed to determine the abundance of AOB on root biofilm samples using fluorescently tagged oligonucleotide probes targeting 16S rRNA. The levels of abundance detected suggested an enrichment of AOB on rice roots. We identified 40 to 69% of AOB on roots of IR63087-1-17 as Nitrosomonas spp., while this subpopulation constituted 7 to 23% of AOB on roots of the other cultivars. These results were generally supported by denaturing gradient gel electrophoresis of the amoA gene and analysis of libraries of cloned amoA. In hydroponic culture, oxygen concentration profiles around secondary roots differed significantly among the tested rice varieties, of which IR63087-1-17 showed maximum leakage of oxygen. The results suggest that varietal differences in the composition and activity of root-associated AOB populations may result from microscale differences in O-2 availability.
  • S Okabe, CM Santegoeds, Y Watanabe, D de Beer BIOTECHNOLOGY AND BIOENGINEERING 78 (2) 119 -130 2002年04月 [査読無し][通常論文]
     
    A combination of fluorescence in situ hybridization (FISH), microprofiles, and denaturing gradient gel electrophoresis (DGGE) analysis of PCR-amplified 16S rDNA fragments followed by hybridization analysis with specific probes was applied to investigate successional development of sulfate-reducing bacteria (SRB) community structure and in situ sulfide production activity within an activated sludge immobilized agar gel film. In this model biofilm system, since biases arising from biofilm heterogeneity can be ignored, the population dynamics of SRB in the agar gel is directly related to physiological capability and in situ activity of SRB. Microelectrode measurements showed that an anoxic zone was already developed at the beginning (0 day), a first sulfide production of 0.054 mumol H2S (m-)2 s(-1) was detected during the first week, and the rate increased gradually to 0.221 mumol H2S m(-2) s(-1) in the fifth week. The most active sulfide production zone moved upward to the chemocline and intensified with time to form a narrow zone with high volumetric sulfide production rates. This result coincided with the shift of the spatial distributions of SRB populations determined by FISH. In situ hybridization with probe SRB385 for mainly general SRB of the delta Proteobacteria plus some gram-positive bacteria and probe 660 for Desulfobulbus indicated that the most abundant populations of SRB were primarily restricted to near the oxic/anoxic interface (chemocline). A close observation of the development of the vertical distributions of SRB populations revealed that the cell numbers of Desulfobulbus tripled (from 0.5 x 10(8) to 1.5 x 108 cells cm(-3)) near the oxic/anoxic interface. Similar growth (from 1.0 x 10(8) to 4.5 x 10(8) cells cm(-3)) of Desulfovibrio-like SRB that hybridized with probe SRB385 was observed. PCR-DGGE followed by hybridization analysis revealed that one Desulfobulbus strain was detected from the beginning, and another strain appeared after 1 week, coinciding with the first detected sulfide production. In addition, three strains hybridizing with probe 687 (possibly Desulfovibrio) were also dominant SRB in the agar gel. (C) 2002 Wiley Periodicals, Inc.
  • 伊藤司, 岡部聡, 渡辺義公, 杉田謙一, 佐藤久 日本水環境学会年会講演集 36th 40 2002年03月14日 [査読無し][通常論文]
  • 金田一智規, 岡部聡, 渡辺義公, 対馬郁夫 日本水環境学会年会講演集 36th 375 2002年03月14日 [査読無し][通常論文]
  • 岸田秀, 岡部聡, 渡辺義公 日本水環境学会年会講演集 36th 345 2002年03月14日 [査読無し][通常論文]
  • 河野快子, 岡部聡, 渡辺義公, 高倉恭子 日本水環境学会年会講演集 36th 374 2002年03月14日 [査読無し][通常論文]
  • T Ito, S Okabe, H Satoh, Y Watanabe APPLIED AND ENVIRONMENTAL MICROBIOLOGY 68 (3) 1392 -1402 2002年03月 [査読無し][通常論文]
     
    A combination of fluorescence in situ hybridization, microprofiles, denaturing gradient gel electrophoresis of PCR-amplified 16S ribosomal DNA fragments, and 16S rRNA gene cloning analysis was applied to investigate successional development of sulfate-reducing bacteria (SRB) community structure and in situ sulfide production activity within a biofilm growing under microaerophilic conditions (dissolved oxygen concentration in the bulk liquid was in the range of 0 to 100 muM) and in the presence of nitrate. Microelectrode measurements showed that oxygen penetrated 200 mum from the surface during all stages of biofilm development. The first sulfide production of 0.32 mumol of H2S m(-2) s(-1) was detected below ca. 500 mum in the 3rd week and then gradually increased to 0.70 mumol H2S m(-2) s(-1) in the 8th week. The most active sulfide production zone moved upward to the oxic-anoxic interface and intensified with time. This result coincided with an increase in SRB populations in the surface layer of the biofilm. The numbers of the probe SRB385- and 660-hybridized SRB populations significantly increased to 7.9 x 10(9) cells cm(-3) and 3.6 x 10(9) cells cm(-3), respectively, in the surface 400 mum during an 8-week cultivation, while those populations were relatively unchanged in the deeper part of the biofilm, probably due to substrate transport limitation. Based on 16S rRNA gene cloning analysis data, clone sequences that related to Desulfomicrobium hypogeium (99% sequence similarity) and Desulfobulbus elongatus (95% sequence similarity) were most frequently found. Different molecular analyses confirmed that Desulfobulbus, Desulfovibrio, and Desulfomicrobium were found to be the numerically important members of SRB in this wastewater biofilm.
  • S Okabe, H Naitoh, H Satoh, Y Watanabe WATER SCIENCE AND TECHNOLOGY 46 (1-2) 233 -241 2002年 [査読無し][通常論文]
     
    The phylogenetic diversity of a nitrifying bacterial community of two types of nitrifying biofilms, a domestic wastewater biofilm and an autotrophic nitrifying biofilm grown on rotating disk reactors (RDR), was characterized by 16S ribosomal DNA (rDNA)-cloning analysis. Thereafter, successional development of nitrifying the bacterial community within both biofilms was visualized in situ by fluorescent in situ hybridization (FISH) with a set of fluorescently labeled 16S rRNA-targeted DNA probes. In situ hybridization revealed that Nitrosomonas ureae was the numerically dominant species of the ammonia-oxidizing population in the domestic wastewater biofilm and that a population shift from N. urea to N. europaea and N. eutropha occurred when the culture medium was switched to the synthetic media from the domestic wastewater. After reaching the steady-state condition, microprofiles of NH4+, NO2-, NO3-, and O-2 in the biofilms were measured by use of microsensors, and the spatial distributions of in situ nitrifying activities were determined. The relationship between the spatial organization of nitrifying bacterial populations and the in situ activity of these populations within the biofilms was discussed. Microelectrode measurements revealed that the active ammonia-oxidizing zone was vertically separated from the active nitrite-oxidizing zone. This vertical separation became more evident with increase of the substrate C/N ratio, leading to deterioration of nitrification efficiency. The combined use of these techniques made it possible to relate in situ nitrifying activity directly to the occurrence of nitrifying bacterial populations.
  • T Ito, JL Nielsen, S Okabe, Y Watanabe, PH Nielsen APPLIED AND ENVIRONMENTAL MICROBIOLOGY 68 (1) 356 -364 2002年01月 [査読無し][通常論文]
     
    We simultaneously determined the phylogenetic identification and substrate uptake patterns of sulfate-reducing bacteria (SRB) inhabiting a sewer biofilm with oxygen, nitrate, or sulfate as an electron acceptor by combining microautoradiography and fluorescent in situ hybridization (MAR-FISH) with family- and genus-specific 16S rRNA probes. The MAR-FISH analysis revealed that Desulfobulbus hybridized with probe 660 was a dominant SRB subgroup in this sewer biofilm, accounting for 23% of the total SRB. Approximately 9 and 27% of Desulfobulbus cells detected with probe 660 could take up [C-14] propionate with oxygen and nitrate, respectively, as an electron acceptor, which might explain the high abundance of this species in various oxic environments. Furthermore, more than 40% of Desulfobulbus cells incorporated acetate under anoxic conditions. SRB were also numerically important members of H-2-utilizing and (CO2)-C-14-fixing microbial populations in this sewer biofilm, accounting for roughly 42% of total H-2-utilizing bacteria hybridized with probe EUB338. A comparative 16S ribosomal DNA analysis revealed that two SRB populations, related to the Desulfomicrobium hypogeium and the Desulfovibrio desulfuricans NIB lineages, were found to be important 11, utilizers in this biofilm. The substrate uptake characteristics of different phylogenetic SRB subgroups were compared with the characteristics described to date. These results provide further insight into the correlation between the 16S rRNA phylogenetic diversity and the physiological diversity of SRB populations inhabiting sewer biofilms.
  • S Okabe, T Kokazi, Y Watanabe 2ND WORLD WATER CONGRESS: WATER DISTRIBUTION AND WATER SERVICES MANAGEMENT 2 (4) 97 -104 2002年 [査読無し][通常論文]
     
    When biodegradable organic matter and other nutrients, such as ammonia and phosphorus, are not sufficiently removed during water treatment, bacteria may proliferate in the water distribution system. Bacterial regrowth deteriorates water quality (taste and odor), accelerates corrosion, and potentially increases the risk of microbial diseases. Therefore, this research was conducted to evaluate the impact of four different advanced water treatment processes, including biological treatments such as a rotating biofilm membrane reactor (RBMR) and a biological activated carbon (BAC) filter and ultrafiltration (UF), on reduction of nutrient levels and biofilm formation potentials of the treated water entering model distribution systems (annular reactors). Our results revealed that biological treatments significantly improved the "biostability" of water leaving from the treatment plant. On average, The RBMR and BAC filter reduced easily assimilable organic carbon (AOC) concentration by half when compared with conventional treatment (multi-media filtration; MF) and ultrafiltration (from 35-49 to 18-23 gg C L-1). Consequently, biofilm formation potential was reduced by a factor of 5 to 10 (from 3,200-5, 100 to 490-710 pg ATP cm(-2)). With respect to "biostability" of water, ultrafiltration was less effective in reducing AOC concentrations. In addition, the impact of chlorine disinfection on biofilm accumulation and AOC levels in the distribution system were studied.
  • S Okabe, H Naitoh, H Satoh, Y Watanabe WATER SCIENCE AND TECHNOLOGY 46 (1-2) 233 -241 2002年 [査読無し][通常論文]
     
    The phylogenetic diversity of a nitrifying bacterial community of two types of nitrifying biofilms, a domestic wastewater biofilm and an autotrophic nitrifying biofilm grown on rotating disk reactors (RDR), was characterized by 16S ribosomal DNA (rDNA)-cloning analysis. Thereafter, successional development of nitrifying the bacterial community within both biofilms was visualized in situ by fluorescent in situ hybridization (FISH) with a set of fluorescently labeled 16S rRNA-targeted DNA probes. In situ hybridization revealed that Nitrosomonas ureae was the numerically dominant species of the ammonia-oxidizing population in the domestic wastewater biofilm and that a population shift from N. urea to N. europaea and N. eutropha occurred when the culture medium was switched to the synthetic media from the domestic wastewater. After reaching the steady-state condition, microprofiles of NH4+, NO2-, NO3-, and O-2 in the biofilms were measured by use of microsensors, and the spatial distributions of in situ nitrifying activities were determined. The relationship between the spatial organization of nitrifying bacterial populations and the in situ activity of these populations within the biofilms was discussed. Microelectrode measurements revealed that the active ammonia-oxidizing zone was vertically separated from the active nitrite-oxidizing zone. This vertical separation became more evident with increase of the substrate C/N ratio, leading to deterioration of nitrification efficiency. The combined use of these techniques made it possible to relate in situ nitrifying activity directly to the occurrence of nitrifying bacterial populations.
  • 伊藤司, 岡部聡, 渡辺義公 日本微生物生態学会講演要旨集 17th 27 2001年11月09日 [査読無し][通常論文]
  • 志水豊晴, 山川岳志, 岡部聡, 渡辺義公 土木学会年次学術講演会講演概要集 第7部 56th 468 -469 2001年09月01日 [査読無し][通常論文]
  • 岡部聡, 渡辺義公, 小鍛冶利彦, 小沢源三 土木学会年次学術講演会講演概要集 第7部 56th 470 -471 2001年09月01日 [査読無し][通常論文]
  • 小鍛冶利彦, 岡部聡, 渡辺義公, 木村克輝 全国水道研究発表会講演集 52nd 278 -279 2001年04月27日 [査読無し][通常論文]
  • 山川岳志, 岡部聡, 渡辺義公 日本水環境学会年会講演集 35th 533 2001年03月14日 [査読無し][通常論文]
  • 志水豊晴, 山川岳志, 岡部聡, 渡辺義公 日本水環境学会年会講演集 35th 318 2001年03月14日 [査読無し][通常論文]
  • S Okabe, H Satoh, Y Watanabe MICROBIAL GROWTH IN BIOFILMS, PT B 337 213 -224 2001年 [査読無し][通常論文]
  • K Kimura, Y Watanabe, S Okabe, H Satoh WATER SCIENCE & TECHNOLOGY: WATER SUPPLY, VOL 1, NO 4 1 (4) 111 -118 2001年 [査読無し][通常論文]
     
    The authors have proposed a novel water treatment process in which nitrifying bacteria are fixed on the surface of rotating membrane disks. This biofilm-membrane process can perform strict solid-liquid separation and oxidation of ammonia nitrogen simultaneously. In this research, applicability of the conventional biofilm model (assuming the biofilm structure to be flat, homogeneous and continuous) to analysis of the biofilm developing in the proposed process was examined. A long-term operation for culturing the active nitrifying biofilm was carried out prior to kinetic investigation. By cryosectioning of the biofilm and image analysis, the thickness of the biofilm was determined to be 87 pm. From this biofilm thickness and the result of the batch ammonia consumption test, the intrinsic zero-order ammonia consumption rate of the biofilm was estimated precisely to be 930 g/m(3)/h. Using these parameters, the ammonia concentration profile in the biofilm was calculated by the conventional model, and the applicability of the model was examined by comparing the calculated profile with the ones measured with a microelectrode. The calculated profile was very close to the measured ones, which indicated feasibility of the conventional model to the analysis of the biofilm grown in the proposed process. The studied biofilm actually had a simple, i.e. flat, homogeneous and continuous, structure due to membrane filtration. This was the reason why the conventional model could still be employed. In the analysis of the data dealing with low concentrations of ammonia, however, first-order kinetics should be used. The first-order ammonia consumption rate constant of the studied biofilm was estimated to be 808 h(-1).
  • 山川岳志, 岡部聡, 渡辺義公 日本微生物生態学会講演要旨集 16th 71 2000年11月12日 [査読無し][通常論文]
  • 伊藤司, 岡部聡, NIELSEN P H, NIELSEN J L, 渡辺義公 日本微生物生態学会講演要旨集 16th 39 2000年11月12日 [査読無し][通常論文]
  • 岡部聡, 渡辺義公, 内藤初夏 土木学会年次学術講演会講演概要集 第7部 55th 122 -123 2000年08月31日 [査読無し][通常論文]
  • 佐藤久, 岡部聡, 渡辺義公 土木学会年次学術講演会講演概要集 第7部 55th 114 -115 2000年08月31日 [査読無し][通常論文]
  • 山川岳志, 岡部聡, 渡辺義公 土木学会年次学術講演会講演概要集 第7部 55th 112 -113 2000年08月31日 [査読無し][通常論文]
  • 山川岳志, 伊藤司, 岡部聡, 渡辺義公 日本水環境学会年会講演集 34th 372 2000年03月15日 [査読無し][通常論文]
  • 佐藤久, 山口有希, 岡部聡, 渡辺義公 日本水環境学会年会講演集 34th 158 2000年03月15日 [査読無し][通常論文]
  • 岡部聡, 渡辺義公 日本水環境学会年会講演集 34th 373 2000年03月15日 [査読無し][通常論文]
  • H Satoh, S Okabe, N Norimatsu, Y Watanabe WATER SCIENCE AND TECHNOLOGY 41 (4-5) 317 -321 2000年 [査読無し][通常論文]
     
    The effect of substrate C/N ratio on the spatial distributions of ammonia-oxidizing bacteria and their activity was investigated by using microelectrodes with high spatial resolution and fluorescent in situ hybridization (FISH) technique. In this study, an interspecies competition for O-2 between ammonia-oxidizing bacteria and heterotrophic bacteria was experimentally evaluated. An autotrophic nitrifying biofilm originally cultured at C/N=0 was used as a model biofilm to study changes in specific NH; oxidation rate profiles in the biofilm when the substrate C/N ratio was varied. As C/N ratio increased, specific NH; oxidation rates decreased in the outer part of the biofilm due to interspecies competition, while they were unchanged in the inner part. The increase in substrate C/N ratio (i.e., addition of acetate) immediately induced the interspecies competition for O-2 between ammonia-oxidizing bacteria and heterotrophic bacteria at the outer part of the biofilm. As a result of the interspecies competition, NH4+ oxidation was restrained, resulting in a decrease in the ammonia-oxidizing bacterial populations. This experimental result clearly explains the stratified spatial distributions of ammonia-oxidizing bacteria within the biofilms at higher substrate C/N ratios. The combined application of microelectrodes and FISH techniques provides new insights into microbial ecology and population dynamics of nitrifying bacteria within multi-species biofilms.
  • S Okabe, Y Watanabe WATER SCIENCE AND TECHNOLOGY 42 (12) 21 -32 2000年 [査読無し][通常論文]
     
    Time dependent development of the spatial organization of NH4+- and NO2--oxidizing bacterial populations in a domestic wastewater biofilm and in an autotrophic nitrifying biofilm were investigated by fluorescent in situ hybridization (FISH) with a set of 16S rRNA-targeted oligonucleotide probes. Population dynamics of nitrifying bacteria in the biofilms were correlated with the biofilm performance. In situ hybridization indicated that Nitrosomonas spp. (excluding probe NEU stained NH4+-oxidizing bacteria: i.e., N. marina-lineage, N, europaea-lineage, N. eutropha, and N. halophila) and Nitrospira-like bacteria were the numerically dominant nitrifying species in the domestic wastewater biofilm. However, probe NEU stained NH4+-oxidizing bacteria became dominant populations in the autotrophic nitrifying biofilm (which were initially cultured with the primary settling tank effluent) after switching to the synthetic media. This population shift might be attributed to the effect of NO2--N accumulation and higher growth Fates of N. europaea-lineage and N. eutropha, outcompeting of her Nitrosomonas spp. in the synthetic medium. This evidence indirectly supports that N. europhaea has been most commonly isolated and studied in most of the previous researches. For the spatial organization of NH4+- and NO2--oxidizing bacterial populations, bacteria of the genus Nitrobacter could not be detected, instead Nitrospira-like bacteria were found as the main nitrite-oxidizing bacteria in both biofilms. Whereas most of the ammonia-oxidizing bacteria were found throughout the biofilms, the location of nitrite-oxidizing bacteria was restricted to the active nitrite-oxidizing zone, which was detected in the inner part of the biofilms. Microelectrode measurements showed that the active ammonia-oxidizing zone was located in the outer part of a biofilm, whereas the active nitrite-oxidizing zone was located just below the ammonia-oxidizing zone and overlapped the location of NO2--oxidizing bacteria, as determined with FISH. These observations have considerable significance to our understanding of microbial nitrification occurring in wastewater treatment processes and in the natural environment.
  • 岡部聡, 佐藤久, 渡辺義公 日本微生物生態学会講演要旨集 15th 36 1999年11月05日 [査読無し][通常論文]
  • S Okabe, T Itoh, H Satoh, Y Watanabe APPLIED AND ENVIRONMENTAL MICROBIOLOGY 65 (11) 5107 -5116 1999年11月 [査読無し][通常論文]
     
    The vertical distribution of sulfate-reducing bacteria (SRB) in aerobic wastewater biofilms grown on rotating dish reactors was investigated by fluorescent in situ hybridization (FISH) with 16S rRNA-targeted oligonucleotide probes. To correlate the vertical distribution of SRB populations with their activity, the microprofiles of O-2, H2S, NO2-, NO3-, NH4+, and pH were measured with microelectrodes. In addition, a cross-evaluation of the FISH and microelectrode analyses was performed by comparing them with culture-based approaches and biogeochemical measurements. In situ hybridization revealed that a relatively high abundance of the probe SRB385-stained cells (approximately 10(9) to 10(10) cells per cm(3) of biofilm) were evenly distributed throughout the biofilm, even in the oxic surface. The probe SRB660-stained Desulfobulbus spp. were found to be numerically important members of SRB populations (approximately 10(8) to 10(9) cells per cm(3)). The result of microelectrode measurements showed that a high sulfate-reducing activity was found in a narrow anaerobic zone located about 150 to 300 mu m below the biofilm surface and above which an intensive sulfide oxidation zone was found. The biogeochemical measurements showed that elemental sulfur (S-0) was an important intermediate of the sulfide reoxidation in such thin wastewater biofilms (approximately 1,500 mu m), which accounted for about 75% of the total S pool in the biofilm. The contribution of an internal Fe-sulfur cycle to the overall sulfur cycle in aerobic wastewater biofilms was insignificant (less than 1%) due to the relatively high sulfate reduction rate.
  • Satoshi Okabe, Tsukasa Itoh, Hisashi Satoh, Yoshimasa Watanabe Applied and Environmental Microbiology 65 (11) 5107 -5116 1999年11月 [査読無し][通常論文]
     
    The vertical distribution of sulfate-reducing bacteria (SRB) in aerobic wastewater biofilms grown on rotating disk reactors was investigated by fluorescent in situ hybridization (FISH) with 16S rRNA-targeted oligonucleotide probes. To correlate the vertical distribution of SRB populations with their activity, the microprofiles of O2, H2S, NO2-, NO3-, NH4+, and pH were measured with microelectrodes. In addition, a cross-evaluation of the FISH and microelectrode analyses was performed by comparing them with culture-based approaches and biogeochemical measurements. In situ hybridization revealed that a relatively high abundance of the probe SRB385-stained cells (approximately 109 to 1010 cells per cm3 of biofilm) were evenly distributed throughout the biofilm, even in the oxic surface. The probe SRB660-stained Desulfobulbus spp. were found to be numerically important members of SRB populations (approximately 108 to 109 cells per cm3). The result of microelectrode measurements showed that a high sulfate- reducing activity was found in a narrow anaerobic zone located about 150 to 300 μm below the biofilm surface and above which an intensive sulfide oxidation zone was found. The biogeochemical measurements showed that elemental sulfur (S0) was an important intermediate of the sulfide reoxidation in such thin wastewater biofilms (approximately 1,500 μm), which accounted for about 75% of the total S pool in the biofilm. The contribution of an internal Fe-sulfur cycle to the overall sulfur cycle in aerobic wastewater biofilms was insignificant (less than 1%) due to the relatively high sulfate reduction rate.
  • 岡部聡, 乗松直生子, 内藤初夏, 渡辺義公 水環境学会誌 22 (8) 683 -691 1999年08月10日 [査読無し][通常論文]
  • 岡部聡, 渡辺義公, 前嶋恵, 黒田浩史 土木学会年次学術講演会講演概要集 第7部 54th 432 -433 1999年08月01日 [査読無し][通常論文]
  • 佐藤久, 岡部聡, 渡辺義公 土木学会年次学術講演会講演概要集 第7部 54th 428 -429 1999年08月01日 [査読無し][通常論文]
  • 木村克輝, 渡辺義公, 岡部聡, 佐藤久 土木学会年次学術講演会講演概要集 第7部 54th 430 -431 1999年08月01日 [査読無し][通常論文]
  • S Okabe, H Satoh, Y Watanabe APPLIED AND ENVIRONMENTAL MICROBIOLOGY 65 (7) 3182 -3191 1999年07月 [査読無し][通常論文]
     
    We investigated the in situ spatial organization of ammonia-oxidizing and nitrite-oxidizing bacteria in domestic wastewater biofilms and autotrophic nitrifying biofilms by using microsensors and fluorescent in situ hybridization (FISH) performed with 16S rRNA-targeted oligonucleotide probes. The combination of these techniques made it possible to relate in situ microbial activity directly to the occurrence of nitrifying bacterial populations. In situ hybridization revealed that bacteria belonging to the genus Nitrosomonas were the numerically dominant ammonia-oxidizing bacteria in both types of biofilms. Bacteria belonging to the genus Nitrobacter were not detected; instead, Nitrospira-like bacteria were the main nitrite-oxidizing bacteria in both types of biofilms. Nitrospira-like cells formed irregularly shaped aggregates consisting of small microcolonies, which clustered around the clusters of ammonia oxidizers. Whereas most of the ammonia-oxidizing bacteria were present throughout the biofilms, the nitrite-oxidizing bacteria were restricted to the active nitrite-oxidizing zones, which were in the inner parts of the biofilms. Microelectrode measurements showed that the active ammonia-oxidizing zone was located in the outer part of a biofilm, whereas the active nitrite-oxidizing zone was located just below the ammonia-oxidizing zone and overlapped the location of nitrite-oxidizing bacteria, as determined by FISH.
  • 佐藤久, 岡部聡, 渡辺義公 日本水環境学会年会講演集 33rd 189 1999年03月15日 [査読無し][通常論文]
  • 山川岳志, 佐藤久, 岡部聡, 渡辺義公 日本水環境学会年会講演集 33rd 191 1999年03月15日 [査読無し][通常論文]
  • 伊藤司, 佐藤久, 岡部聡, 渡辺義公 日本水環境学会年会講演集 33rd 190 1999年03月15日 [査読無し][通常論文]
  • FISH法を用いた硝化細菌生物膜内の生態学的構造解析
    水環境学会誌 22 (8) 683 -691 1999年 [査読無し][通常論文]
  • DR Noguera, S Okabe, C Picioreanu WATER SCIENCE AND TECHNOLOGY 39 (7) 273 -278 1999年 [査読無し][通常論文]
     
    Biofilm models are commonly used as simulation tools in engineering applications and as research tools to identify and fill gaps in our knowledge of biofilm processes. While models used in engineering applications rely on simplifying assumptions to make them practical, recent experimental evidence of biofilm heterogeneity questions the validity of these assumptions. On the other band, research models are becoming more complex and use advanced computational tools to mathematically investigate which factors determine the structural heterogeneity and the population dynamics of biofilms. One of the goals of advanced models is to evaluate the relevance of three-dimensional heterogeneities to the predictive capability of traditional biofilm models. In addition, biofilm models are used to evaluate experimental observations when studying a diversity of biofilm-related phenomena. Given the variety of applications of biofilm models and the different approaches that modelers have taken in recent years, a specialist group was convened to evaluate the present status and determine future directions of biofilm modeling research. The education of scientists and engineers on the fundamentals of biofilm models, the development of mathematical models for real-time control of biofilm processes, and the ability to "engineer" the biofilm structure and function (or performance) were identified as the most important objectives for the practical application of biofilm models. As mathematical research tools, biofilm models are directed towards gaining a better understanding of biofilm structure and population dynamics. Specific topics identified as priorities on biofilm research include the behavior of specialist microorganisms, the elucidation of attachment and detachment mechanisms, the determination of mechanical properties of exopolymeric substances, and the study of ecological interactions among different microorganisms. The need to evaluate parameter sensitivity in the different models was identified as an essential component of modeling research. A group decision from this meeting was to initiate a collaborative effort to identify similarities and differences among current modeling approaches. Such comparative analysis will enhance our understanding of biofilm processes and mathematical approaches, and will facilitate the future use of biofilm models by scientists and engineers involved in biofilm research. (C) 1999 IAWQ Published by Elsevier Science Ltd. All rights reserved.
  • S Okabe, H Satoh, T Itoh, Y Watanabe WATER SCIENCE AND TECHNOLOGY 39 (7) 41 -47 1999年 [査読無し][通常論文]
     
    The vertical distribution of sulfate-reducing bacteria (SRB) in microaerophilic wastewater biofilms grown on fully submerged rotating disk reactors (RDR) was determined by the conventional culture-dependent MPN method and in situ hybridization of fluorescently-labelled 16S rRNA-targeted oligonucleotide probes for SRB in parallel. Chemical concentration profiles within the biofilm were also measured using microelectrodes for O-2, S2-, NO3- and pH. In situ hybridization revealed that the SRB probe-stained cells were distributed throughout the biofilm even in the oxic surface zone in ail states from single scattered cells to clustered cells. The higher fluorescence intensity and abundance of SRB probe-stained cells were found in the middle part of the biofilm. This result corresponded well with O-2 and H2S concentration profiles measured by microelectrodes, showing sulfate reduction was restricted to a narrow anaerobic zone located about 500 mu m below the biofilm surface. Results of the MPN and potential sulfate reducing activity (culture-dependent approaches) indicated a similar distribution of cultivable SRB in the biofilm, The majority of the general SRB probe-stained cells were hybridized with SRB 660 probe, suggesting that one important member of the SRB in the wastewater biofilm could be the genus Desulfobulbus. An addition of nitrate forced the sulfate reduction zone deeper in the biofilm and reduced the specific sulfate reduction rate as well. The sulfate reduction zone was consequently separated from O-2 and NO3- respiration zones. Anaerobic H2S oxidation with NO3- was also induced by addition of nitrate to the medium.
  • 佐藤久, 岡部聡, 渡辺義公 水環境学会誌 22 (3) 206 -214 1999年 [査読無し][通常論文]
  • 岡部聡, 内藤初夏, 渡辺義公 水環境学会誌 22 (3) 191 -198 1999年 [査読無し][通常論文]
  • 佐藤久, 岡部聡, 渡辺義公 水環境学会誌 22 (9) 763 -769 1999年 [査読無し][通常論文]
  • 伊藤司, 佐藤久, 岡部聡, 渡辺義公 環境工学研究フォーラム講演集 35th 147 -149 1998年11月 [査読無し][通常論文]
  • 岡部聡, 乗松直生子, 渡辺義公 環境工学研究フォーラム講演集 35th 162 -164 1998年11月 [査読無し][通常論文]
  • 内藤初夏, 佐藤久, 岡部聡, 渡辺義公, 乗松直生子 土木学会年次学術講演会講演概要集 第7部 53rd 152 -153 1998年10月 [査読無し][通常論文]
  • 佐藤久, 岡部聡, 渡辺義公, 乗松直生子 土木学会年次学術講演会講演概要集 第7部 53rd 154 -155 1998年10月 [査読無し][通常論文]
  • 岡部聡, 渡辺義公, 川村美穂 土木学会年次学術講演会講演概要集 第7部 53rd 158 -159 1998年10月 [査読無し][通常論文]
  • 大西徳子, 伊藤司, 岡部聡, 渡辺義公 土木学会年次学術講演会講演概要集 第7部 53rd 156 -157 1998年10月 [査読無し][通常論文]
  • 渡辺義公, 岡部聡, 鈴木辰彦 マンガン自触媒反応と嫌気性膜洗浄を利用した膜処理の効率化 平成9年度 No.08555135 6,7-23 1998年 [査読無し][通常論文]
  • 渡辺義公, 岡部聡, 鈴木辰彦 マンガン自触媒反応と嫌気性膜洗浄を利用した膜処理の効率化 平成9年度 No.08555135 24,25-49 1998年 [査読無し][通常論文]
  • AOCを指標とした高度浄水処理システムの性能評価
    水道協会雑誌 67 (11) 12 -21 1998年 [査読無し][通常論文]
  • 水環境学会誌 21 (2) 88 -97 1998年 [査読無し][通常論文]
  • 佐藤久, 岡部聡, 伊藤司, 渡辺義公 水環境学会誌 21 (6) 367 -375 1998年 [査読無し][通常論文]
  • S Okabe, T Matsuda, H Satoh, T Itoh, Y Watanabe WATER SCIENCE AND TECHNOLOGY 37 (4-5) 131 -138 1998年 [査読無し][通常論文]
     
    The microzonation of O-2 respiration, H2S oxidation and SO42- reduction in aerobic biofilms grown on rotating disk reactors of a sewage treatment plant was studied by measuring concentration profiles with microelectrodes for O-2, S2- and pH. The vertical distribution of sulfate-reducing bacteria (SRB) in the biofilms was also determined by the conventional culture-dependent MPN method and fluorescently labeled 16S rRNA-targeted oligonucleotide probes for SRB. The SRB probe stained cells were distributed throughout the biofilm with a distinct higher fluorescence intensity in the middle part of the biofilm. This result corresponded well with O-2 and S2- concentration gradients measured by microelectrodes, showing sulfate reducing activity was largely restricted to a narrow anaerobic zone located in the middle of the biofilm. Measurements of accumulation of reduced sulfur compounds (FeS, FeS2 and S-O) in the biofilm indicated that the H2S produced by SRB became oxidized by O-2 and other oxidants, probably ferric/ferrous hydrates, and precipitated as FeS and S-O just above the sulfate reduction zone. (C) 1998 IAWQ. Published by Elsevier Science Ltd.
  • S Okabe, H Kuroda, Y Watanabe WATER SCIENCE AND TECHNOLOGY 38 (8-9) 163 -170 1998年 [査読無し][通常論文]
     
    Evolutional changes in interior structures of mixed population biofilms grown on domestic wastewater were quantitatively analyzed using a cryosectioning technique and an image analysis. Meanwhile, transport of particulates into the biofilms was also experimentally investigated using fluorescent microbeads as tracers to relate the biofilm structure and particulate transport into the biofilm. Microscopic observation of the cryomicrotomy biofilm sections indicated the biofilms were very porous and consisted of interwinded filamentous biomass acting as a framework of the biofilm. A honeycomb structure was often found, which would make the biofilm more resistant to water flow. There were micropores with the diameter of about 10 mu m microcolony aggregates attached to filamentous biomass and macropores with the diameter of 20-200 mu m in the biomass matrix. These pores did not clog during two months of cultivation. Areal porosity was about 30% in the bottom biofilm and more than 90 % in the surface. Significant difference in transport efficiency was not observed for various sizes of microbeads due to the presence of macropores. Therefore, even 10 mu m tracer beads could quickly traverse throughout a biofilm 640 mu m thick via water channels or macropores and then penetrated into the micropores. Convective transport from the bulk to the bottom biofilm, rather than molecular diffusion, was responsible for this rapid transport. Based on experimental results, it can be concluded that the biofilm structure seems to be well designed to maximize the transport efficiency of substrates and products and the strength of biofilm structure. (C) 1998 IAWQ, Published by Elsevier Science Ltd. All rights reserved.
  • 伊藤司, 乗松直生子, 佐藤久, 岡部聡, 渡辺義公, 境一澄 環境工学研究フォーラム講演集 34th 91 -93 1997年11月 [査読無し][通常論文]
  • 佐藤久, 岡部聡, 伊藤司, 渡辺義公, 松田尚之 環境工学研究フォーラム講演集 34th 94 -96 1997年11月 [査読無し][通常論文]
  • 佐藤久, 岡部聡, 渡辺義公, 境一澄 土木学会年次学術講演会講演概要集 第7部 52nd 294 -295 1997年09月 [査読無し][通常論文]
  • 岡部聡, 渡辺義公, 境一澄 土木学会年次学術講演会講演概要集 第7部 52nd 296 -297 1997年09月 [査読無し][通常論文]
  • 岡部聡, 佐藤久, 渡辺義公, 尾原裕昌 土木学会年次学術講演会講演概要集 第7部 52nd 330 -331 1997年09月 [査読無し][通常論文]
  • 岡部聡, 乗松直生子, 境一澄, 渡辺義公, 船田良 日本水環境学会年会講演集 31st 135 1997年03月 [査読無し][通常論文]
  • 黒田浩史, 岡部聡, 渡辺義公 日本水環境学会年会講演集 31st 134 1997年03月 [査読無し][通常論文]
  • 佐藤久, 伊藤司, 岡部聡, 渡辺義公 日本水環境学会年会講演集 31st 136 1997年03月 [査読無し][通常論文]
  • 境一澄, 岡部聡, 渡辺義公 日本水環境学会年会講演集 31st 133 1997年03月 [査読無し][通常論文]
  • S Okabe, T Yasuda, Y Watanabe BIOTECHNOLOGY AND BIOENGINEERING 53 (5) 459 -469 1997年03月 [査読無し][通常論文]
     
    Inert fluorescent microparticles were used as tracers to investigate the dynamics of spatial distribution of particulate components in mixed population biofilms. The tracer bead spatial distributions in the biofilm were experimentally measured by sectioning the biofilms with a microslicer. The experimental results were compared with model simulations using the biofilm model (BIOSIM) to evaluate the assumption that advective transport (displacement) of particulates balances with cell growth in the model. The tracer beads could traverse throughout a biofilm 360 mu m thick with in less than 23 minutes, which cannot be explained solely by their attachment to the surface followed by molecular diffusion. Advective transport of the tracer beads via ''voids and pores'' could be responsible for such rapid bead penetration. Observation by confocal scanning laser microscopy (CSLM) clearly showed that the biofilm consisted of a thick loose surface layer, varying in thickness, and a semicontiguous base layer separated by water channels. About 80% of attached tracer beads remained in the biofilm for over 20 days. The trapped tracer beads were gradually transferred from the depth of the biofilm to the surface. The observed bead release rate was much slower than the model predictions. This is probably because the cell density increased predominantly near the substratum, resulting in an unbalance of advective transport of the tracer beads and cell growth. The pores, voids, and cell-free spaces in the biofilm were first filled with growing biomass, thereafter, displacement of the beads took place once the cell density reached certain levels. The model assumptions of the temporal and spatial constant cell density and the continuum concept (flat biomass) are clearly oversimplified and should be revised. It was concluded that the dynamics of the inert microbeads in the biofilm was strongly influenced by not only microbial growth, but also by the biofilm structure and growth pattern. Therefore, one dimensional modeling is not adequate for the accurate description of the transport of particulates in a biofilm. (C) 1997 John Wiley & Sons, Inc.
  • Y Watanabe, K Kimura, S Okabe, G Ozawa, N Ohkuma WATER SCIENCE AND TECHNOLOGY 36 (1) 51 -60 1997年 [査読無し][通常論文]
     
    For the oxidation of low concentrations of NH4+-N, conventional biofilm reactors such as a rotating biological contactor encounter difficulty due to mass transport limitation of NH4+-N. Therefore, the authors have developed a novel biofilm-membrane reactor, in which biomass is fixed on the surface of rotating membrane disks to enhance NH4+-N transport into the biofilm. Three long-term bench-scale experiments were carried out and sufficient nitrification efficiency was obtained even at low levels of NH4+-N. The experimental results were evaluated in comparison with model simulation. (C) 1997 IAWQ. Published by Elsevier Science Ltd.
  • 木質系および石炭系粒状活性炭を用いたオゾン・生物活性炭処理
    水道協会雑誌 66 (12) 20 -29 1997年 [査読無し][通常論文]
  • S Okabe, K Hirata, Y Watanabe BIOFOULING 11 (2) 119 -136 1997年 [査読無し][通常論文]
     
    Dynamic changes in spatial microbial distribution in mixed-population biofilms resulting from interspecies competition between heterotrophs and nitrifiers were experimentally investigated using a microslicer technique. Biofilms cultured in partially submerged rotating biological contactors (RBC) with synthetic wastewater were used as test materials. The results showed that variation in the carbon loading rate (CLR) and the composition of the feed substrate resulted in rapid and substantial changes in the spatial distribution of nitrifiers in mixed population biofilms within a week, which significantly influenced the NH4-N removal rate. Heterotrophs were more successful than nitrifiers in acquiring dissolved oxygen and space and dominated throughout the biofilms, especially in the surface biofilm. Nitrifiers were therefore diluted in the surface biofilm and mainly found in the inner biofilm. The fraction differences of NH4-oxidizers and NO2-oxidizers over the biofilm depth were about 2 orders and 3-4 orders of magnitude at CLR=0.8 and 1.6 g-C m(-2) d(-1) respectively, indicating a steep spatial gradient of nitrifiers. Although total areal density of NH4-oxidizers in the biofilms were in the same order, the NH4-N flux decreased by about 9% and 24% after one month at CLR=0.8 and 1.6 g-C m(-2) d(-1) respectively. This decrease of NH4-N flux was attributed to the extent of the stratified spatial distribution of NH4-oxidizers. Heterotrophs lie near the surface and act as a diffusion barrier, resulting in an increase in internal dissolved oxygen and NH4-N diffusion resistance for NH4-oxidizers. These results suggest that simplifying or neglecting the spatial distribution of microbial species can lead to substantial errors in biofilm kinetic parameters determined from measured substrate removal rates. For implication in reactor design, the extent of the spatial distribution of microbial species in a biofilm is especially important for interpretation of nitrification efficiency in the presence of organic matter.
  • 笠原伸介, 相沢拓, 渡辺義公, 小沢源三, 岡部聡, 丹保憲仁 土木学会年次学術講演会講演概要集 第7部 51st 212 -213 1996年08月 [査読無し][通常論文]
  • 乗松直生子, 岡部聡, 渡辺義公 土木学会年次学術講演会講演概要集 第7部 51st 340 -341 1996年08月 [査読無し][通常論文]
  • 佐藤久, 黒田浩史, 岡部聡, 渡辺義公 土木学会年次学術講演会講演概要集 第7部 51st 102 -103 1996年08月 [査読無し][通常論文]
  • 井野場誠治, 岡部聡, 渡辺義公 土木学会年次学術講演会講演概要集 第7部 51st 342 -343 1996年08月 [査読無し][通常論文]
  • 岡部聡, 渡辺義公, 安田岳雄 土木学会年次学術講演会講演概要集 第7部 51st 112 -113 1996年08月 [査読無し][通常論文]
  • S Okabe, K Hiratia, Y Ozawa, Y Watanabe BIOTECHNOLOGY AND BIOENGINEERING 50 (1) 24 -35 1996年04月 [査読無し][通常論文]
     
    Spatial microbial distributions of nitrifiers and heterotrophs in undefined mixed-population biofilms were experimentally investigated using a microslicer technique and correlated with nitrification efficiency of the biofilm system. The general stratification of different bacterial groups in the biofilm was simulated using a one-dimensional (1-D) mathematical biofilm accumulation model (BAM) and compared with the experimental results. Biofilms were cultured at three C:N ratios of feed solutions In a partially submerged rotating biological contactor (RBC). It was shown that the biofilms were vertically stratified (from biofilm surface to substratum). At C:N = 0, heterotrophs and nitrifiers coexisted in the outermost biofilm and heterotrophs dominated in the innermost biofilm. At C:N = 1.5, heterotrophs outcompeted nitrifiers for dissolved oxygen and space; thus, heterotrophs dominated in the outermost biofilm and nitrifiers were present only in the deeper biofilm. Nitrifiers and heterotrophs coexisted in the innermost biofilm. An increase in the influent C:N ratio resulted in stronger stratification of microbial species, as well as inhibition of nitrification. In batch experiments, NH4-N utilization rate (R(NH4-N)) was almost the same at each substrate C: N ratio even though NH, oxidizers were predominantly present in the deeper biofilm. The biofilm performance could not be sufficiently explained by the obtained microbial spatial distribution, suggesting that one-dimensional description of microbial distribution was not good enough and three-dimensional measurements of microbial spatial distribution is necessary. Total bacterial densities increased by a factor of 3-17 with biofilm depth. The metabolically active cell fraction decreased from 35 +/- 13% in the outermost biofilm to 15 +/- 4% in the innermost biofilm, presumably due to substrate limitation. The model predicted more pronounced stratification of nitrifiers and heterotrophs than the observed results. This discrepancy could be attributed to the real biofilms that were structurally heterogeneous (e.g., water channels), which could not be described by the one-dimensional model. The results of this study clearly indicate the limitation of 1-D biofilm models to describe the extent of stratification of nitrifiers and heterotrophs and suggest a 3-D model is necessary. (C) 1996 John Wiley & Sons, Inc.
  • 松田尚之, 岡部聡, 渡辺義公 日本水環境学会年会講演集 30th 131 1996年03月 [査読無し][通常論文]
  • S. Okabe, Y. Oozawa, K. Hirata, Y. Watanabe Water Research 30 (7) 1563 -1572 1996年 [査読無し][通常論文]
     
    The relationship between time dependent population dynamics of nitrifiers and heterotrophs in undefined mixed-population biofilms and their nitrification efficiency was experimentally investigated at various C:N ratios of feed solutions. Five types of biofilms were cultured in partially submerged rotating biological contactors (RBC's) at different C:N ratios and were used as test materials. The results indicated that initial microbial composition in the biofilms and substrate composition (e.g. C:N ratio) strongly influenced the later population dynamics and the nitrification efficiency. Higher influent C:N ratio retarded accumulation of nitrifying bacteria, especially NO2-oxidizers, resulting in a considerably long start-up period for complete and stable nitrification due to competition for dissolved oxygen and space in the biofilm. Furthermore, a start-up inoculum was very important to keep start-up time of nitrification to a minimum. Time-dependent population dynamics in the biofilms reflected well the bulk water quality and microbial community structure in the bulk liquid. These results suggest that the structure of microbial community in the biofilm can be predicted from monitoring the water quality and microbiology of the bulk liquid. Physiologically inactive cells in the biofilm were determined by an INT dehydrogenease assay. These cells gradually accumulated up to about 30% of the total bacterial population within the biofilms. The results of this study will provide a rational basis for developing and controlling desired biofilm population dynamics to maximize nitrification efficiency of wastewater biofilms.
  • 混合培養系生物膜内の懸濁微粒子の挙動に関する基礎的研究
    環境工学研究論文集 33 103 -114 1996年 [査読無し][通常論文]
  • 嫌気化した茨戸湖底泥の硫黄循環を中心とした生物化学的現状把握
    環境工学研究論文集 33 331 -340 1996年 [査読無し][通常論文]
  • 岡部聡, 渡辺義公, 境一澄 土木学会年次学術講演会講演概要集 第2部 50th (B) 1142 -1143 1995年09月 [査読無し][通常論文]
  • OKABE S, NIELSEN P H, JONES W L, CHARACKLIS W G Water Research 29 (2) 571 -578 1995年01月01日 [査読無し][通常論文]
     
    Sulfide product inhibition kinetics for growth and activity of Desulfovibrio desulfuricans was investigated in batch and continuous cultures at pH = 7.0. A non-competitive inhibition model adequately described sulfide product inhibition kinetics. Inhibition coefficient (Ki) for maximum specific growth rate (μinhmax) was 251 mg l-1S in a batch experiment. Cell yield determined in a chemostat was reduced in half by a sulfide concentration of about 250 mg l-1S, which was very close to the Kivalue for the batch growth. Maximum specific growth rate (μinhmax) and cell yield (Yc Lac) were strongly inhibited by high levels of sulfide concentrations, whereas specific lactate utilization rate increased with increasing sulfide concentrations. The results indicated an increase in the relative energy needed for maintenance to overcome sulfide inhibition and uncoupling growth from energy production. However, D. desulfuricans to some extent could recover from the shock of high sulfide concentrations. Stoichiometry for catabolic reactions (energy producing) did not change at high sulfide concentrations, while anabolic reactions (cellular synthesis) were strongly inhibited by high sulfide concentrations. These results suggested that separation of sulfide product inhibition into growth (cell yield) and activity (substrate utilization rate) was important to incorporate the sulfide product inhibition kinetics in a variety of applications. © 1995.
  • Y WATANABE, S OKABE, K HIRATA, S MASUDA WATER SCIENCE AND TECHNOLOGY 31 (1) 195 -203 1995年 [査読無し][通常論文]
     
    Simultaneous nitrification with denitrification was investigated using a single rotating biological contactor (RBC). The authors proposed two means to achieve simultaneous nitrification with denitrification in the single reactor: (1) to control oxygen transfer rate by reducing oxygen partial pressure (P-O2) in the air phase and (2) to develop a combined partially (aerobic) and fully (anaerobic) submerged RBC (CPFSR) reactor. For the former experiment, the maximum denitrification efficiency of 90 % was obtained at C:N ratio=6 and P-O2=0.10 atm. Moreover, heterotrophs, NH4- and NO2- oxidizers, and denitrifiers were distributed throughout the biofilm, suggesting that nitrification and denitrification can occur wheresoever the local environment meets their growth conditions. For the latter experiment, effects of type of organic matter and influent carbon:nitrogen ratio (C:N ratio) on the efficiency of simultaneous nitrification with denitrification were investigated using the CPFSR reactor. Acetate, ethylene-glycol, phenol, and poly-vinyl-alcohol (PVA) were used as carbon sources for denitrification. An excellent nitrification efficiency was obtained for all experimental runs and all organic substrates could be degraded and used for denitrification, indicating a great potential for simultaneous removals of nitrogen and xenobiotic compounds by the CPFSR reactor.
  • S OKABE, PH NIELSEN, WL JONES, WG CHARACKLIS BIOFOULING 9 (1) 63 -83 1995年 [査読無し][通常論文]
     
    The kinetics and stoichiometry of Desulfovibrio desulfuricans attached to a polycarbonate surface were determined in RotoTorque(TM) reactors and compared with those of suspended cells. Biofilm specific cellular growth rate (mu(b)) and detachment rate (q(dx)) were determined from unsteady state biofilm experiments. In the initial biofilm accumulation phase, the specific cellular growth rate was the same as the maximum specific growth rate for D. desulfuricans in suspension (mu(max) = 0.37 . h(-1)); thereafter mu(b) decreased and approached a steady state value of about 0.1 . h(-1). The decrease in average cellular specific growth rate could be attributed to substrate (lactate) limitation in some experiments, but in others there was no evidence of this. Biofilm-specific cellular detachment rate decreased similarly to biofilm-specific cellular growth rate. In biofilms, cellular yield at mu(b) = 0.1 . h(-1) was approximately 18% of planktonic cellular yield partly due to the production of extensive extracellular polymeric substance. A linear relationship between mu(b) and specific lactate utilization rate (q(s)) in the biofilm did not exist. During the steady state biofilm accumulation phase (mu(b) = .0.1 . h(-1)), specific lactate utilization by biofilm cells was about 2-3 times greater than by planktonic cells, whereas it was essentially the same during the initial biofilm accumulation phase. These results suggest that kinetic and stoichiometric data derived from suspended cells must be cautiously incorporated into biofilm accumulation models.
  • S Okabe, K Hirata, Y Watanabe WATER SCIENCE AND TECHNOLOGY 32 (8) 67 -74 1995年 [査読無し][通常論文]
     
    Dynamic changes in spatial microbial distribution in mixed-population biofilms were experimentally determined using a microslicer technique and simulated by a biofilm accumulation model (BAM). Experimental results were compared with the model simulation. The biofilms cultured in partially submerged rotating biological contactors (RBC) with synthetic wastewater were used as test materials. Experimental results showed that an increase of substrate loading rate (i.e., organic carbon and NH4-N) resulted In the microbial stratification in the biofilms. Heterotrophs defeated nitrifiers and dominated in the outer biofilm, whereas nitrifiers were diluted out in the outer biofilm and forced into the inner biofilm. At higher organic loading rates, a stronger stratified microbial spatial distribution was observed, which imposed a severe internal oxygen diffusion limitation cn nitrifiers and resulted in the deterioration of nitrification efficiency. Model simulations described a general trend of the stratified biofilm structure. However, the actual stratification was stronger than the simulated results. For implication in the reactor design, when the specific carbon loading rate exceeds a certain limit, nitrification will be deteriorated or require a long start-up period due to the interspecies competition resulting in oxygen diffusion limitation. The extend of microbial stratification in the biofilm is especially important for determination of feasibility of nitrification in the presence of organic matters.
  • 岡部聡, 渡辺義公 環境工学研究フォーラム講演集 31st 124 -126 1994年11月 [査読無し][通常論文]
  • 岡部聡, 渡辺義公, 平田貴紅子 土木学会年次学術講演会講演概要集 第2部 49th (B) 1074 -1075 1994年09月 [査読無し][通常論文]
  • 渡辺義公, 岩崎義彦, 岡部聡 水文・水資源学会研究発表会要旨集 1994 368 -369 1994年08月 [査読無し][通常論文]
  • 岡部聡, 渡辺義公, 大沢祐子 土木学会西部支部研究発表会講演概要集 1993 312 -313 1994年03月 [査読無し][通常論文]
  • 三木幸浩, 岡部聡, 渡辺義公 土木学会西部支部研究発表会講演概要集 1993 310 -311 1994年03月 [査読無し][通常論文]
  • Y WATANABE, S OKABE, T ARATA, Y HARUTA WATER SCIENCE AND TECHNOLOGY 30 (11) 25 -33 1994年 [査読無し][通常論文]
     
    A comprehensive wastewater treatment system that accomplishes oxidation of organic matter, nitrification, and denitrification was developed, and its characteristics and performance were investigated. A municipal wastewater was treated by an up-flow aerated biofilter (UAB), in which biofilms were developed on stainless meshes installed horizontally. This UAB exhibited a great potential ability of oxidation of organic matter, SS stabilization, and nitrification due to a unique aeration mechanism giving high DO concentrations with relatively low aeration rates. Another unique feature of the UAB was that attached biofilms on stainless meshes physically filtered out and/or adsorbed suspended solids in the wastewater in addition to the biological oxidation of organic matter. A stable nitrification could be achieved at HRT = 10 hours corresponding to a hydraulic loading of 86 L m(-2) d(-1) and at a ratio of aeration rate to wastewater flow rate (A/W) of 2, which is considerably low as compared to aeration rates of typical activated sludge systems. This UAB system also could handle relatively high hydraulic loading rates. The UAB used in this study still have enough space to install more stainless meshes so as to reduce hydraulic loading rates resulting in the reduction of HRT and aeration rate, which leads to improvement of the system performance as well as reduction of the running cost.
  • OKABE S, NIELSEN P H, JONES W L, CHARACKLIS W G . Water Research 28 (11) 2263 -2266 1994年01月01日 [査読無し][通常論文]
     
    Measurement of cellular and extracellular carbon contents of sulfate-reducing bacteria (SRB) is essential and important in studies of the role of SRB in corrosion and biofouling. An epifluorescence (EPI) microscopic technique and a lipopolysaccharide (LPS) assay were used to quantify cellular and extracellular carbon contents in Desulfovibrio desulfuricans biofilms. The average contents of lipopolysaccharide (LPS) and cellular carbon were 7.3 ± 2.8 (fg LPS) cell-1 and 39.9 ± 9.9 (fg cellular-C) cell-1, respectively, in a D. desulfuricans chemostat culture. A ratio of cellular carbon content to LPS content was 6.5 ± 2.8, and was used to estimate cellular carbon contents in a D. desulfuricans biofilm. The LPS and EPI methods gave comparable results for suspended samples, but not for biofilm samples. © 1994.
  • Y WATANABE, S OKABE, T ARATA, Y HARUTA WATER SCIENCE AND TECHNOLOGY 30 (11) 25 -33 1994年 [査読無し][通常論文]
     
    A comprehensive wastewater treatment system that accomplishes oxidation of organic matter, nitrification, and denitrification was developed, and its characteristics and performance were investigated. A municipal wastewater was treated by an up-flow aerated biofilter (UAB), in which biofilms were developed on stainless meshes installed horizontally. This UAB exhibited a great potential ability of oxidation of organic matter, SS stabilization, and nitrification due to a unique aeration mechanism giving high DO concentrations with relatively low aeration rates. Another unique feature of the UAB was that attached biofilms on stainless meshes physically filtered out and/or adsorbed suspended solids in the wastewater in addition to the biological oxidation of organic matter. A stable nitrification could be achieved at HRT = 10 hours corresponding to a hydraulic loading of 86 L m(-2) d(-1) and at a ratio of aeration rate to wastewater flow rate (A/W) of 2, which is considerably low as compared to aeration rates of typical activated sludge systems. This UAB system also could handle relatively high hydraulic loading rates. The UAB used in this study still have enough space to install more stainless meshes so as to reduce hydraulic loading rates resulting in the reduction of HRT and aeration rate, which leads to improvement of the system performance as well as reduction of the running cost.
  • Whonchee Lee, Zbigniew Lewandowski, Sathoshi Okabe, William G. Characklis, Recep Avci Biofouling 7 1 -15 1993年11月01日 [査読無し][通常論文]
     
    The sulfate-reducing bacteria (SRB)-enhanced corrosion of mild steel in the presence of 1.5 mg.l-1 dissolved oxygen (DO) in bulk liquid was investigated. The biofilm process analysis was combined with microelectrode measurements, electrochemical measurements, and surface analysis. In the early stages of biofilm accumulation, the cathodic polarization and the decreasing corrosion rate were attributed to DO consumption by aerobic bacteria. During that time, limited SRB activity was observed. The DO concentration near the steel surface was between 0.6 and 1 mg.l-1. After total depletion of dissolved oxygen near the steel surface, the cathodic depolarization and the increased corrosion rate were associated with the proliferation of SRB near the steel surface. Auger electron spectroscopy analysis indicated localized sulfide attack. High pit density appeared where the coincidence of oxygen and sulfur occurred. The bottom of the pit was enriched with sulfur. © 1993, Taylor & Francis Group, LLC. All rights reserved.
  • 岡部聡, 渡辺義公, 春田勇二, 荒田朋睦 土木学会年次学術講演会講演概要集 第2部 48th 1312 -1313 1993年09月 [査読無し][通常論文]
  • Anaerobic biofilms in industrial water-systems
    Abstracts of Papers of the American Chemical Society 204 9999 1993年 [査読無し][通常論文]
  • OKABE S, CHARACKLIS W G Biotechnology and Bioengineering 39 (10) 1031 -1042 1992年01月01日 [査読無し][通常論文]
     
    The effects of temperature and phosphorous concentration on the rate and the extent of microbial sulfate reduction with lactate as carbon and energy source were investigated for Desulfovibrio desulfuricans. The continuous culture experiments (chemostat) were conducted at pH 7.0 from 12 to 48°C. The maximum specific growth rate (μmax) was relatively constant in the range 25°C–43°C and dramatically decreased outside this temperature range. The half‐saturation coefficient was minimum at 25°C. Cell yield was highest in the optimum temperature range (35°C–43°C) for growth. Maintenance energy requirements for D. desulfuricans were not significant. Two moles of lactate is consumed for every mole of sulfate reduced, and this stoichiometric ratio is not temperature dependent. Steady state rate and stoichiometric coefficients accurately predicted transient behavior during temperature shifts. The extent of extracellular polymeric substance (EPS) is related to the concentration of phosphorous in the medium. EPS production rate increased with decreased phosphorous loading rate. Failure to discriminate between cell and EPS formation by D. desulfuricans leads to significant overestimates of the cell yield. The limiting C:P ratio for D. desulfuricans was in the range of 400:1 to 800:1. Copyright © 1992 John Wiley & Sons, Inc.
  • Corrosion of mild steel underneath aerobic biofilms containing sulfate-reducing bacteria
    Corrosion 47 1 -15 1992年 [査読無し][通常論文]
  • OKABE S, NIELSEN P H, CHARACKLIS W G Biotechnology and Bioengineering 40 (6) 725 -734 1992年01月01日 [査読無し][通常論文]
     
    The effects of sulfate and nitrogen concentrations of the rate and stoichiometry of microbial sulfate reduction were investigated for Desulfovibrio desulfuricans grown on lactate and sulfate in a chemostat at pH 7.0. Maximum specific growth rates (μmax), half‐saturation coefficients (Ksul), and cell yield (Yc/Lac) of 0.344 ± 0.007 and 0.352 ± 0.003 h−1, 1.8 ± 0.3 and 1.0 ± 0.2 mg/L, and 0.020 ± 0.003 and 0.017 ± 0.003 g cell/g lactate, respectively, were obtained under sulfate‐limiting conditions at 35°C and 43°C. Maintenance energy requirements for D. desulfuricans were significant under sulfate‐limiting conditions. The extent of extracellular polymeric substance (EPS) produced was related to the carbon: nitrogen ratio in the medium. EPS production rate increased with decreased nitrogen loading rate. Nitrogen starvation also resulted in decreased cell size of D. desulfuricans. The limiting C : N ratio (w/w) for D. desulfuricans was in the range of 45 : 1 to 120 : 1. Effects of sulfide on microbial sulfate reduction, cell size, and biomass production were also ivestigated at pH 7.0. Fifty percent inhibition of lactate utilization occurred at a total sulfide concentration of approximately 500 mg/L. The cell size of D. desulfuricans decreased with increasing total sulfide concentration. Sulfide inhibition of D. desulfuricans was observed to be a reversible process. © 1992 John Wiley & Sons, Inc. Copyright © 1992 John Wiley & Sons, Inc.
  • 渡辺義公, 石黒政儀, 岡部聡, 田代雄児 土木学会年次学術講演会講演概要集 第2部 41st 807 -808 1986年11月 [査読無し][通常論文]

書籍等出版物

  • Nitrification
    ASM Press 2011年
  • バイオ燃料電池の最新動向
    シーエムシー出版 2011年
  • Research on Nitrification and Related Processes, Part B
    Elsevier 2011年
  • Nitrification
    ASM Press 2011年
  • Recent treands in microbial fuel cells
    CMC 2011年
  • Research on Nitrification and Related Processes, Part B
    Elsevier 2011年
  • 水再利用学
    技報堂出版 2010年
  • 難培養微生物研究の最新技術II
    シーエムシー出版 2010年
  • Wastewater Treatment
    Caister Academic Press 2010年
  • Water Reuse
    2010年
  • Wastewater Treatment
    Caister Academic Press 2010年

所属学協会

  • 国際微生物生態学会   International Water Association   「Environmental Technology」編集委員   「Biodegradation」編集委員   American Society for Microbiology   土木学会   「Microbes and Environments」副編集委員長   「Water Research」Associate Editor   「Water Science and Technology」Associate Editor   日本微生物生態学会   International society for microbial ecology   

共同研究・競争的資金等の研究課題

  • バイオ燃料電池駆動型エネルギー自立式Anammox MECシステムの開発
    日本学術振興会:科学研究費助成事業 基盤研究(A)
    研究期間 : 2019年04月 -2023年03月 
    代表者 : 岡部 聡, 北島 正章, 佐藤 久, 押木 守
     
    嫌気性アンモニア酸化(Anammox)プロセスは、省エネ型窒素除去プロセスとして工場排水や嫌気性消化脱水ろ液処理などに適用が始まっているが、メインストリームの都市下水処理への適用は一向に進んでいない。メインストリームの都市下水処理への適用の鍵は、前段の部分硝化(NH4+をNO2-まで酸化する)プロセスの安定化・高効率化である。 本研究では、前段に有機物除去を担うバイオ燃料電池(MFC)を設け、MFCで発生する電圧を用いてMECアノード電極電位を制御することで、アンモニア酸化細菌(AOB)がアノード電極を電子受容体としてNH4+をNO2-まで酸化し、Anammox細菌が生成されたNO2-とNH4+を窒素ガス(N2)へ変換するMFC駆動型部分硝化(PN)-Anammox 生物電解セル(MEC)システムを開発するものである。本年度は以下の2点について検討した。① MFCを補助電源とし、アンモニア酸化細菌(AOB)は、アノード電極を電子受容体としてNH4+をNO2-へ酸化できるか?さらに、Anammox細菌は、アノード電極を電子受容体としてNH4+を直接N2へ酸化できるか? ② 亜硝酸酸化細菌(NOB; NO2-をNO3-に酸化する細菌)に特異的に感染し溶菌させるバクテリオファージは存在するのか?そして、それをNOB増殖抑制剤として利用し部分硝化反応(NH4+ → NO2-)の安定化を効率的に達成できるか?その結果、①関してはMFCを補助電源とするMECを構築し、NH4+の除去性能を確認できた。②に関しては、活性汚泥などからファージを濃縮しNitrospiraのバイオマスと混合し、ファージ感染の有無を確認する実験を行ったが、NOBに特異的に感染するバクテリオファージを獲得するには至らなかった。
  • 非生物・生物ハイブリッド人工光合成システムの構築:持続可能な酢酸生成拠点の創出
    日本学術振興会:科学研究費助成事業 挑戦的研究(開拓)
    研究期間 : 2019年06月 -2022年03月 
    代表者 : 岡部 聡, 渡辺 精一, 佐藤 久
  • 日本学術振興会:科学研究費助成事業 基盤研究(A)
    研究期間 : 2012年04月 -2015年03月 
    代表者 : 大村 達夫, 原田 秀樹, 佐野 大輔, 渡部 徹, 岡部 聡, 真砂 佳史
     
    本研究では、水環境中におけるノロウイルスの未知動態を解明することを目的とし、1)ウイルス外殻タンパク質損傷評価法の適用条件、2)環境由来濃縮サンプルから目的ウイルスゲノムのみを特異的に回収する手法の開発、および3)ノロウイルスによる養殖カキの汚染度に関する調査・研究を行った。その結果、カプシドタンパク質損傷検出手法が水環境中のヒトノロウイルスに適用可能であることが確認され、環境由来濃縮サンプルから損傷を受けていないウイルスゲノムのみを特異的に回収する手法の確立に成功した。さらに養殖カキ中の優占している遺伝子型のみならずマイナーな感染流行株が次世代シーケンスにより検出可能であることが示された。
  • 糞便汚染マーカー定量検出を基盤とした微生物学的水質管理手法の確立
    日本学術振興会:科学研究費助成事業 基盤研究(A)
    研究期間 : 2011年04月 -2014年03月 
    代表者 : 岡部 聡, 石井 聡, 佐藤 久, 佐野 大輔
     
    現行の糞便汚染指標である大腸菌群数による微生物学的水質管理は、水環境中での大腸菌群の増殖、糞便に由来しない大腸菌群の存在など、数多くの問題点が指摘されている。このため、合理的な糞便性汚染指標の確立および水域の微生物学的水質管理手法の確立が急務となっている。このような背景のもと、本研究では、宿主特異的遺伝子マーカー(腸内蛋白質分解細菌の最優占種であるBacteroides-Prevotella 属由来遺伝子)をもとに水域の糞便汚染レベルを定量的に評価し、さらに糞便汚染源(ヒト、家畜及び野生動物等)を迅速かつ正確に特定する新規方法論を確立し、具体的な汚染防止対策の構築を含む合理的な微生物学的水質管理を実現することを目的としている。 本年度は、昨年度開発した各宿主特異的糞便汚染マーカーを実際の水環境に適用し、糞便汚染の実態を明らかにした。さらに、遺伝子マーカーを糞便汚染指標として活用するためには、生存細胞と死細胞を区別して定量することが求められる。そこで、Propidium monoazide (PMA)を併用した定量PCR法を確立し、糞便汚染源の特定を行うために重要となる生菌由来の糞便性汚染マーカーの定量が可能となった。 次に、環境水中における糞便汚染マーカー(ヒト、ブタ、ウシ、ニワトリ、カモの各宿主特異的遺伝子マーカー)の挙動を解析した。さらに、既存の糞便汚染指標である大腸菌群数、糞便生大腸菌群及び大腸菌の定量も行い、減衰速度の違いを評価した。 最後に、水系感染する腸管系感染症起因細菌及びノロウイルス等の腸管系ウイルスの特異的検出を行い、各宿主特異的遺伝子マーカーと病原微生物の環境水中における存在比の相関関係を調査した。
  • 水循環の基盤となる革新的水処理システムの創出
    JST戦略的創造研究推進制度(研究チーム型) (戦略的基礎研究推進事業:CREST)
    研究期間 : 2009年 -2014年
  • Development of innovative water and wastewater treatment systems for sustainable urban water metabolism
    JST Basic Research Programs (Core Research for Evolutional Science and Technology :CREST)
    研究期間 : 2009年 -2014年
  • 日本学術振興会:科学研究費助成事業 挑戦的萌芽研究
    研究期間 : 2011年 -2012年 
    代表者 : 岡部 聡
     
    本研究は、廃水処理と電気エネルギー回収が同時に可能となることが期待されているバイオ燃料電池のさらなる発電能力の向上を図ることを目的とした。そこで本年度は、カソード反応を促進するため、すなわちプロトン供給量を促進するために、カソードへ供給する空気中の水分およびCO2濃度の発電量に及ぼす影響について検討した。供給する空気中のCO2濃度が高くなるほど、カソード電極表面上に存在する高pH水に溶解するCO2が増大しプロトンの供給を促し、電気伝導率の向上が確認された。さらに、供給するガス組成(CO2とO2の濃度比)および供給速度を変化させて、最大の電力密度が得られる条件を求めた。
  • トキシコゲノミクスによる新規機能性ナノ高分子(デンドリマー)の毒性評価
    日本学術振興会:科学研究費助成事業 挑戦的萌芽研究
    研究期間 : 2009年 -2010年 
    代表者 : 岡部 聡
     
    PAMAM(polyamidoamine)デンドリマーの生物学的活性は世代が大きくなる程高くなるため、毒性は世代に強く依存すると考えられる。世代依存毒性を含むPAMAMデンドリマーの毒性のより詳細な解明が必須であり、本研究ではPAMAMデンドリマーの毒性及びその発現メカニズムの解明を目的とした。ヒト肝癌由来細胞株HepG2細胞を用いて、PAMAMデンドリマーG4-G7の毒性評価を行った。全ての試験通して、PAMAMデンドリマーとBSAの相互作用による影響を除く為、PAMAMデンドリマーの溶媒にHBSS(Hank's balanced salt solution)にHEPESを10mMとなるように加えたものを用いた。また、暴露濃度にはNeutral red(NR)assayで細胞生存率が80%となる濃度をそれぞれ用いた。その結果、PAMAMデンドリマーの世代依存的な細胞毒性を確認した。同じNR assayで細胞生存率が80%となる濃度でも、世代の大きいPAMAMデンドリマーほどROSの産生能が高いことが明らかとなった。ROS産生は120minで最高値をとった。さらに、DNA損傷性を評価するために、H_2O_2をPositive Controlとしてコメットアッセイを行った。実験結果の解析は、尾の長さ、面積、濃さにより、尾の無いもの(I)から,核内のDNA全てが断片化した頭部の無いものあるいは頭部の極微少なもの(V)までI~Vの5段階に分けて評価した。その結果、AMAMデンドリマーで処理していないHBSS(negative control)の細胞群では、クラスIが大部分であった(75.2%)のに対し、PAMAMデンドリマー暴露系ではクラスIが40%~55%と低く、G7で最低値の40.4%%となった。また、G6,G7では損傷の重篤であるIV,Vの割合が高く、コメット値はG5を除いて世代が増加するに従い、増加する傾向が認められた。以上の結果より、PAMAMデンドリマーがDNA損傷性を有しており、DNA損傷の程度は世代が大きい程より重篤であることが明らかとなった。また、1mM H_2O_2とPAMAMデンドリマーG6はROS産生量がほぼ同程度であったにも関わらず、DNA損傷レベルはH_2O_2の方がPAMAMデンドリマーに比べかなり高い値となった。このことから、酸化ストレスがPAMAMデンドリマーの毒性の直接的な原因ではない可能性が示唆された。
  • 日本学術振興会:科学研究費助成事業 基盤研究(B)
    研究期間 : 2008年 -2010年 
    代表者 : 岡部 聡
     
    本研究では、ヒトDNAマイクロアレイを用いて、代表的な汚染物質である重金属、農薬の混合暴露下におけるヒト由来細胞(HepG2)の遺伝子発現解析を行い、重金属、農薬それぞれの暴露で得られた遺伝子発現パターンと比較し複合毒性作用(相乗、拮抗、もしくは新たな毒性の出現)の解明および有用な毒性マーカー遺伝子の選定を行った。さらに、遺伝子発現解析により選定された遺伝子マーカーを用いて、より詳細な遺伝子発現解析(定量的RT-PCR)を実施し、各重金属-農薬の組合せにおいて、遺伝子発現レベルで作用濃度閾値を明らかにした。
  • ヒトDNAマイクロアレイを用いたナノ粒子の細胞毒性評価
    日本学術振興会:科学研究費助成事業 萌芽研究
    研究期間 : 2007年 -2008年 
    代表者 : 岡部 聡
     
    現在生産されているナノ粒子の毒性に関する情報は限られており、人体への影響は解明されてはいない。本研究では、この技術を銀ナノ粒子存在下ヒト由来細胞に応用し、銀ナノ粒子の毒性の有無およびその毒性作用について考察を行った。ニュートラルレッド法と形態観察の結果より、遺伝子発現解析を実施する暴露濃度を低濃度でありながら形態変化が明らかであった40μg/Lと、より高濃度の1000μg/Lに決定した。対照系で紡錘形であった細胞が、銀ナノ粒子暴露群では仮足が伸びたような形態へと変化し、細胞内に多数の空隙が認められた。データベースを基に有意変動遺伝子の機能分類を行った結果、銀、ポリスチレン両ナノ粒子暴露においてM-phaseに分類され、細胞分裂を促進する作用を持つ遺伝子群の上昇変動が顕著であった。また、DNA-repairに分類される遺伝子群の上昇変動も顕著であった。これは、DNAの修復と対応していると考えられ、遺伝子傷害の可能性が示唆された。また、銀ナノ粒子暴露に特異的な遺伝子発現パターンとして、活性酸素種の解毒に重要な役割を担う遺伝子をはじめとする酸化ストレス応答遺伝子の下降変動が挙げられた。小核試験の結果、銀ナノ粒子暴露系にのみ多くの小核が確認され(50.2%)、銀ナノ粒子暴露による遺伝子障害の可能性を強く支持していると考えられる。また、アスコルビン酸を蓄積させた細胞で同様の実験を行った結果、小核形成率が低下したことから(50.2%→26.3%)、銀ナノ粒子暴露による遺伝子障害性に、活性酸素種の関与が示唆された。以上の結果をまとめると、銀、ポリスチレンナノ粒子において発癌作用に関連すると推測される細胞増殖刺激、遺伝子障害を示唆する遺伝子発現パターンが確認された。さらに、これらの結果は、小核試験によって支持され、また、遺伝子傷害性がアスコルビン酸によって抑制されることから、銀ナノ粒子暴露による遺伝子障害性に対する活性酸素種の関与が示唆された。
  • DNAマイクロアレイを用いた環境汚染化学物質の多指標型毒性評価システムの開発
    日本学術振興会:科学研究費助成事業 基盤研究(B)
    研究期間 : 2005年 -2007年 
    代表者 : 岡部 聡
     
    本年度は、発癌作用のモデル化学物質として、ニトロソアミン(DNA障害性)、TPA(DNA非障害性、プロモーター)、TCE(DNA非障害性)を選択し、これら化学物質により特異的に誘導される遺伝子ライブラリーを構築した。また、3種の重金属(As、 Cd、 Ni)暴露における遺伝子発現解析を行い、モデル化学物質暴露における遺伝子発現パターンと比較した。その結果、これら重金属の主要な発癌作用は細胞増殖の促進、DNA傷害であることが明らかとなった。 上記の特徴は活性酸素(ROS)産生物質であるDMNQでも認められたため、ROS産生が重金属の主要な発癌メカニズムと仮定し、抗酸化物質であるアスコルビン酸処理後の細胞についても同様の検討を行った。その結果、ヒ素では細胞増殖、DNA傷害に関わる変動遺伝子数が顕著に減少したのに対し、カドミウムとニッケルではむしろ増加する結果となった。すなわち、ヒ素の発癌性においては活性酸素産生が主要な発癌メカニズムであり、カドミウム、ニッケルにおいては他のメカニズムが関与していることが明らかとなった。 さらに本研究では、発癌性マーカーとして有用であると考えられる遺伝子PTTG1が見出され、供試したすべての発癌物質、重金属において有意に発現することが定量的RT-PCR法によって確認された。重金属の発癌性はエイムズテスト等の既存の発癌評価本手法によって評価することが困難であるため、簡便かつ迅速な評価手法の開発が求められる。本研究によって得られた結果より、DNAマイクロアレイによって重金属を含む広範囲の発癌物質を評価可能であることが明らかとなり、また、見いだされた発癌性マーカーPTTG1遺伝子は機構の異なる発癌物質の包括的なスクリーニングに有用であると考えられる。
  • 遺伝子および化学マーカーによる河川糞便性汚染源の特定
    日本学術振興会:科学研究費助成事業 基盤研究(B)
    研究期間 : 2003年 -2004年 
    代表者 : 岡部 聡, 木村 克輝
     
    本研究では糞便性指標微生物の遺伝子マーカーと化学マーカーを組み合わせた迅速(1-2日程度)かつ簡便な、糞便性汚染源の特定と汚染度の定量的評価手法の開発を行うことを目的とした。腸内蛋白質分解細菌の最優占種であるBacteroidesに着目し、培養を必要としない分子生物学手法(16S rDNAクローンライブラリー法やTerminal Restriction Fragment Length Polymorphism : T-RFLP法、定量PCR法)を用い、宿主特有のBacteroidesの16S rRNA遺伝子配列(遺伝子マーカー)を特定・探索することにより、水環境中の糞便汚染源(人間、牛、豚等)を特定する。研究成果は以下のように要約される。 宿主動物(人間、牛、豚)毎に特異的なBacteroides種が存在し、宿主動物を認識することが可能な16SrRNA遺伝子配列(遺伝子マーカー)を見つけ出すことに成功した。これによって、これら特異的なBacteroides属を指標微生物とすることにより、糞便汚染源の特定が可能であることを示唆している。次に、これら宿主特異的遺伝子マーカーを迅速かつ簡便に検出・定量するために、これらの遺伝子マーカーに特異的なPCRプラーマーのセットを7つ設計することができた。これら設計したPCRプラーマーセットを用いたT-RFLP法を確立した。この結果、宿主動物毎に特異的なDNA断片長を有するクローンが河川環境中に多く存在することが確認された。したがって、これらのDNA断片は宿主動物毎の遺伝子マーカーとなることが確認できた。また、遺伝子マーカーの迅速な定量を行うために、それぞれのプライマーセットに対応するReal-timePCR法の反応条件を確立した。T-RFLP法およびReal-timePCR法による解析は、DNA抽出から定量まで約8時間で終了し、かつ培養法によるバイアスを排除できるため、より正確に複合微生物系内に存在する各種糞便性大腸菌および病原細菌の特定・定量および汚染源の特定・モニタリングが可能となると思われる。また、検出限界は極めて低く、高感度かつ特異的な糞便性大腸菌汚染の指標となることが明らかとなった。
  • 再生水造水とリン資源回収のためのハイブリッド下水処理システムの構築
    日本学術振興会:科学研究費助成事業 基盤研究(A)
    研究期間 : 2002年 -2004年 
    代表者 : 渡辺 義公, 岡部 聡, 木村 克輝
     
    1)膜ファウリング機構の解明および抑制 実都市下水を用いた多系列の膜分離活性汚泥処理実験を行い、運転条件、前処理の有無が反応槽内汚泥の性状に及ぼす影響について検討した。反応槽内汚泥濃度の上昇に伴って槽内汚泥の粘度が上昇し、膜ろ過運転を困難にするが、特に汚泥濃度が10,000mg/Lを超過する際に粘度の上昇が顕著になる傾向が観察された。前凝集沈殿処理を行うことでこれらの粘度上昇は良好に抑制されうることを見いだした。 2)ハイブリッドMBRでは窒素除去率が低いことを改善するために、PVDF製のMF平膜を用いたMBR内に仕切り板を入れて、MBRの水位を変化させて槽内に好気部と無酸素部を同時に形成する新たなMBRを開発した。このMBRでは処理水の全窒素濃度が5mg/l程度となった。また、MBR内の生物相の解析によって、MBR内には糸状細菌であるChloroflexiが活性汚泥ばっき槽内と比較して極めて高い存在割合を占めていることを明らかにした。Chloroflexiは死滅細菌由来の有機物のスカベンジャーとして機能し、溶解性有機物の蓄積による膜ファウリングの抑制に寄与していることが示唆された。 3)処理水の再利用の観点から、Bacteroides属とPrevotella属を指標として、迅速かつ正確に人獣の糞便由来の病原性微生物の存在をチェックできる新たな分子生物学的手法を開発し、実河川の水質測定に適用した。その結果開発された手法によって、人、牛、豚の糞便汚染を区別できることが明らかになり、処理水の安全性の確認のみではなく、河川の糞便汚染の汚染源を特定できるようになった。 4)新規吸着剤であるジルコニアメゾ構造体(ZS)をリン酸吸着に適用した結果、ZSに含まれるSO_4^<2->とOH^-がリン酸イオンと陰イオン交換することで、リン酸を選択的に吸着する機構を明らかにした。その最大吸着能約3500μM/gZSであり、既存の吸着剤の1.7倍に達した。ZSに吸着されたリン酸はクエン酸溶液(pH=4.5)やNaOH溶液に浸すことで容易に溶出した。今後はZSの再生方法と高濃度・高純度に濃縮されたリン酸溶液からの硝析脱リンによるリン回収技術の確立を計画している。
  • バイオ燃料電池の開発
    研究期間 : 2004年
  • Development of microbial fuel cells
    研究期間 : 2004年
  • 世界の環境改善という視点から見た環境工学の未来とその発展のための政策提案
    日本学術振興会:科学研究費助成事業 基盤研究(C)
    研究期間 : 2003年 -2003年 
    代表者 : 福士 謙介, 飯田 俊彰, 岡部 聡, 大瀧 雅寛, 滝沢 智, 徳永 光晴
     
    我が国の環境工学はこれまで一定の成果をあげてきた。そのおかげで、国民は衛生的かつ快適な生活を送り、世界の最長寿国として知られるようになった。また、生物が住めないような極度に汚染された河川・湖沼・海洋は日本中に存在しない。その一方で、目を東南アジア、南アジア、アフリカ、南アメリカなどに向けると、環境の崩壊は確実に進みつつある。 本研究は世界に目を向けた場合、開発が必要である環境工学の諸分野とそのタスクを明らかにすることを第一の目的とした。次に、それらの研究開発(技術移転を含む)を効率的に行い、内外の研究者が開発に専一に取り組むための政府や助成団体ならびの大学の研究助成体制のあり方や総合的援助システムを提言することを第二の目的とした。最終的には本研究理念を具現化するための研究グループを立ち上げる為の活動を具体的に行う提案をする事を活動の最終的な目的とした。 本研究活動として平成15年8月に北海道イトムカにおいて第1回ワークショップを合宿形式で開催した。約20名の若手研究者が集まり、次世代の環境工学について協議を行った。次に、タイ・バンコク市において外国の研究者を交えて第2回ワークショップを開催した。その結果平成16年度、文部科学省・科学技術振興調整費・我が国の国際的リーダーシップの確保のプログラムへ「アジアの持続的発展のための国際協議会企画」というプロジェクトの提案を行った。この提案課題は中堅〜若手の柔軟性のある研究者や実務者が参集し長時間にわたり、持続的な環境について協議する場を提案するもので、最終的にはアジアにおける、環境を中心テーマとした、いわゆるダボス会議(世界経済フォーラム年次総会)を目指す物である。
  • バイオフィルムの形成機構に関する研究
    研究期間 : 2000年
  • トキシコゲノミクスを用いた化学物質の毒性評価手法の開発
    研究期間 : 2000年
  • Study on biofilm development
    研究期間 : 2000年
  • Evaluation of chemical toxicity by using toxicogenomics
    研究期間 : 2000年
  • 生物膜の機能と生態学的構造に関する研究
  • Biofilm structure and function


Copyright © MEDIA FUSION Co.,Ltd. All rights reserved.