Molecular characterization and diversity of thermophilic iron-reducing enrichment cultures from deep subsurface environments

AIMS: The objectives of this work were to explore the diversity in Fe (III)-reducing enrichment cultures from the deep subsurface and to identify strains involved in metal reduction. METHODS AND RESULTS: Analyses of 16S ribosomal RNA (rRNA) of enrichments, supplemented with hydrogen, acetate or pyruvate as an electron donor, identified three dominant operational taxonomic units (OTUs). All cultures exhibited considerable diversity (36-24 OTUs), even after being transferred at least nine times. Two OTUs were present in all three cultures, constituting about 65% of the total clones examined. CONCLUSION: Dominant OTUs appeared to be most closely related to Thermoanaerobacter ethanolicus or T. kivui. One OTU, which is potentially responsible for autotrophic Fe (III) reduction, was only about 95% similar to T. ethanolicus and may represent a new species. SIGNIFICANCE AND IMPACT OF THE STUDY: An unexpectedly high diversity was found in these enrichments and this diversity may be a feature that can be exploited.     

Diversity of Culturable Bacteria,from the Anaerobic Zone of the Meromictic Lake Pavin,Able to Perform Dissimilatory-Iron Reduction in Different in Vitro Conditions

A culture-dependent study was performed with the aim of assessing the carbon, electron and Fe(III) sources used for the dissimilatory Fe(III) reduction pathway and the diversity of culturable Fe(III)-reducers in the anoxic zone of the meromictic Lake Pavin. This metabolic pathway was investigated in enrichment cultures inoculated with water samples collected at 70 m depth in the anoxic zone of Lake Pavin. Combinations of different media, organic acids, and incubation gas phases were performed. The potential for Fe(III) reduction in the different growth conditions was assessed by measuring the accumulation of Fe(II) overtime. Bacterial community structure was determined in each growth conditions by Temporal Temperature gradient Gel Electrophoresis (TTGE) profiles of 16S rDNA genes and bands of interest in positive enrichments were sequenced. Comparisons of bacterial community structure between growth conditions revealed that the electron donor, the basal media as well as the Fe(III) source yielded to the selection of different bacterial populations, suggesting that Fe(III) reducers occupy different ecological niches in the anoxic zone of Lake Pavin. Facultative Fe(III) reducers, such as fermentative (e.g., Pseudomonas, Clostridium) and sulphate-reducing (e.g., Desulfovibrio sp.) bacteria, were retrieved in enrichments but well-known obligatory Fe(III) reducers (e.g., Geobacter) were not detected. A greater Fe(III) reduction was noted under H₂:CO₂ gas phase, suggesting that H₂ is used as an electron donor for Fe(III) reduction. Acetate was not used as a precursor for this terminal electron-accepting process, and a high Fe(III) reduction was observed with fumarate provided as the electron donor and carbon sources suggesting that this metabolite may be energetically more beneficial for Fe(III)-reducers.     

温州市境内城市河流底泥氨氧化菌富集培养物微生物群落结构分析

【背景】城市河流底泥含有丰富的微生物资源,底泥表面更是硝化作用的主要位点之一,其表面微生物在河流生态系统氮的转化过程中发挥着重要作用。【目的】以温州市境内的城市河流水系温瑞塘河茶山段舜岙河和横江河的4条河道作为采样点,比较分析4种不同环境下城市河流表层底泥氨氧化菌富集培养物的微生物群落结构。【方法】通过野外采样及室内培养对底泥中氨氧化功能菌进行富集培养,采用高通量测序技术分析微生物群落的组成、丰度和多样性。【结果】富集培养后主要优势类群为变形菌门(Proteobacteria)和拟杆菌门(Bacteroidetes)。4个样品共涉及氨氧化细菌3个属,分别为亚硝化单胞菌属(Nitrosomonas)、亚硝化螺菌属(Nitrosospira)、亚硝化球菌属(Nitrosococcus),涉及氨氧化古菌1个属为Nitrososphaera,其中所有样品均以Nitrosomonas为主。不同底泥富集样品氨氧化微生物可操作分类单元(Operational taxonomic unit,OTU)组成存在明显差异,栽种有水生植物的河道底泥样品DA2具有最高的氨氧化细菌OTU数量和相对丰度,而存在生活餐饮污染的河道底泥样品DA4具有最高的氨氧化古菌OTU数量和相对丰度;相较于滞留水体,采自相对流动水体的富集样品DA2、DA4具有更高的氨氧化微生物OTU数量和相对丰度。【结论】阐述了4种不同环境下城市河流底泥氨氧化菌富集培养物微生物群落结构的多样性,确定了富集培养之后的优势类群,为氨氧化微生物培养源的选择提供了参考,也为城市河流底泥中氨氧化菌进一步的筛选分离及其生理生态特征的研究提供了科学依据。     

Bacterial diversity in the bulk soil and rhizosphere fractions of Lolium perenne and Trifolium repens as revealed by PCR restriction analysis of 16S rDNA

The rhizosphere of Trifolium repens and Lolium perenne was divided into three fractions: the bulk soil, the soil adhering to the roots and the washed roots (rhizoplane and endorhizosphere). After isolation and purification of DNA from these fractions, 16S rDNA was amplified by PCR and cloned to obtain a collection of 16S rRNA genes representative of the bacterial communities of these three fractions. The genes were then characterized by PCR restriction analysis. Each different profile was used to define an operational taxonomic unit (OTU). The numbers of OTUs and the numbers of clones among these OTUs allowed to calculate a diversity index. The number of OTUs decreased as root proximity increased and a few OTUs became dominant, resulting in a lower diversity index. In the root fraction of T. repens, the restriction profile of the dominant OTU matched the theoretical profile of the 16S rRNA gene of Rhizobium leguminosarum. This study showed that plant roots create a selective environment for microbial populations.     

REVIEWS

Atlas of Mapped Distributions of Dominance and Modern Pollen Percentages for Important Tree Taxa of Eastern North America . By P. A. D elcourt , H. R. D elcourt and T. W ebb III.
New Manual of Bryology. Volume 1. Edited by R. M. S chuster .
The Ecology and Physiology of the Fungal Mycelium. Edited by D. H. J ennings and A. D. M. R ayner .
Isotopes and Radiation in Agricultural Sciences. Edited by M. E. L'A nnunziata and J. O. L egg .
The Physiology of Flowering Plants : their growth and development . Third edition. By H. E. S treet and H. Ö pik .
Reconstructing Quaternary Environments. By J. J. L owe and M. J. C. W alker .
European Mires. Edited by P. M oore .
Compartments in Algal Cells and Their Interaction. (Proceedings in Life Sciences Series.) Edited by W. W iessner , D. G. R obinson and R. C. S tarr .
Chloroplast Metabolism. By B arry H alliwell .
Flooding and Plant Growth. Ed. by T. T. K ozlowski .
Mode of Action of Antifungal Agents. Ed. by A. P. J. T rinci and J. F. R yley .
Disease Resistance in Plants. By J. E. V anderplank .     

Phylogenetic and physiological diversity of dissimilatory ferric iron reducers in sediments of the polluted Scheldt estuary, Northwest Europe

The potential for dissimilatory ferric iron [Fe(III)] reduction in intertidal sediments of the polluted Scheldt estuary, Northwest Europe, was assessed by combining field-based geochemical measurements with laboratory experiments on the associated microbiology. Microbial communities at a freshwater and brackish location were characterized by culture-independent 16S rRNA gene analysis, as well as enrichments, strain isolation and physiological screening. Dilution-to-extinction batch enrichments using a variety of Fe(III) sources were performed. The dilution factor of the inoculum in the enrichments had a more determining effect on the Fe(III)-reducing microbial community structure than the Fe(III) source. Well-known Fe(III) reducers, including members of the family Geobacteraceae and the genus Shewanella, constituted only a small fraction (< or = 1%) of the in situ microbial community. Instead, facultative anaerobic Ralstonia and strictly anaerobic, spore-forming Clostridium species dominated Fe(III) reduction. These species were able to utilize a variety of electron acceptors. This flexibility may help the organisms to survive in the dynamic estuarine environment. The high diversity and abundance of culturable Fe(III) reducers (4.6 x 10(5) and 2.4 x 10(4) cells g(-1) sediment at the freshwater and brackish site respectively), plus the high concentrations of chemically reducible solid-phase Fe(III) at the sites, implied a high potential for dissimilatory Fe(III) reduction in the estuarine sediments. Pore water chemical data further supported in situ dissimilatory Fe(III) reduction.     

Comparison of Humic Substance- and Fe(III)-Reducing Microbial Communities in Anoxic Aquifers

Humic substances can mediate electron transfer between microorganisms and Fe(III) minerals. Because it is unknown which microorganisms reduce humics in anoxic aquifers, we analyzed the diversity and physiological flexibility of Fe(III)-, humics-, and AQDS-reducers, which were present at up to 10⁶ cells g^?1. No significant differences in 16S rRNA gene based diversity were found between enrichment cultures reducing ferrihydrite, humics or AQDS. Even after repeated transfers many of the enrichments retained the ability to switch to other electron acceptors. This suggests that humics- and Fe(III)-reducing microorganisms in anoxic aquifers are rather versatile and able to reduce different extracellular electron acceptors.     

Reviews

Book Reviewed in this article:
Fortschritte der Chemie organischer Naturstoffe: Progress in the Chemistry of Organic Natural Products , vol. 51. Founded by L. Z echmkister . Ed. by W. H erz , H. G risebach , G. W. K irby and C. T amm .
Immobilized Cells: Principles and Applications. By J. T ampion and M. D. T ampion .
Cell and Tissue Culture in Forestry, Volume 3. Case Histories: Gymnosperms, Angiosperms and Palms. Ed. by J. M. B onga and D. J. D urzan .
Anatomy of the Dicotyledons. Second Edition. Vol III. Magnoliales, Illiciales, and Law ales. By C. R. M etcalfe .
The Ecology of the Nitrogen Cycle. By J anet I. S prent .     

Enumeration and characterization of iron(III)-reducing microbial communities from acidic subsurface sediments contaminated with uranium(VI)

Iron(III)-reducing bacteria have been demonstrated to rapidly catalyze the reduction and immobilization of uranium(VI) from contaminated subsurface sediments. Thus, these organisms may aid in the development of bioremediation strategies for uranium contamination, which is prevalent in acidic subsurface sediments at U.S. government facilities. Iron(III)-reducing enrichment cultures were initiated from pristine and contaminated (high in uranium, nitrate; low pH) subsurface sediments at pH 7 and pH 4 to 5. Enumeration of Fe(III)-reducing bacteria yielded cell counts of up to 240 cells ml(-1) for the contaminated and background sediments at both pHs with a range of different carbon sources (glycerol, acetate, lactate, and glucose). In enrichments where nitrate contamination was removed from the sediment by washing, MPN counts of Fe(III)-reducing bacteria increased substantially. Sediments of lower pH typically yielded lower counts of Fe(III)-reducing bacteria in lactate- and acetate-amended enrichments, but higher counts were observed when glucose was used as an electron donor in acidic enrichments. Phylogenetic analysis of 16S rRNA gene sequences extracted from the highest positive MPN dilutions revealed that the predominant members of Fe(III)-reducing consortia from background sediments were closely related to members of the Geobacteraceae family, whereas a recently characterized Fe(III) reducer (Anaeromyxobacter sp.) and organisms not previously shown to reduce Fe(III) (Paenibacillus and Brevibacillus spp.) predominated in the Fe(III)-reducing consortia of contaminated sediments. Analysis of enrichment cultures by terminal restriction fragment length polymorphism (T-RFLP) strongly supported the cloning and sequencing results. Dominant members of the Fe(III)-reducing consortia were observed to be stable over several enrichment culture transfers by T-RFLP in conjunction with measurements of Fe(III) reduction activity and carbon substrate utilization. Enrichment cultures from contaminated sites were also shown to rapidly reduce millimolar amounts of U(VI) in comparison to killed controls. With DNA extracted directly from subsurface sediments, quantitative analysis of 16S rRNA gene sequences with MPN-PCR indicated that Geobacteraceae sequences were more abundant in pristine compared to contaminated environments,whereas Anaeromyxobacter sequences were more abundant in contaminated sediments. Thus, results from a combination of cultivation-based and cultivation-independent approaches indicate that the abundance/community composition of Fe(III)-reducing consortia in subsurface sediments is dependent upon geochemical parameters (pH, nitrate concentration) and that microorganisms capable of producing spores (gram positive) or spore-like bodies (Anaeromyxobacter) were representative of acidic subsurface environments.     

Multiple approaches to enhance the cultivability of bacteria associated with the marine sponge Haliclona (gellius) sp

Three methods were examined to cultivate bacteria associated with the marine sponge Haliclona (gellius) sp.: agar plate cultures, liquid cultures, and floating filter cultures. A variety of oligotrophic media were employed, including media with aqueous and organic sponge extracts, bacterial signal molecules, and siderophores. More than 3,900 isolates were analyzed, and 205 operational taxonomic units (OTUs) were identified. Media containing low concentrations of mucin or a mixture of peptone and starch were most successful for the isolation of diversity, while the commonly used marine broth did not result in a high diversity among isolates. The addition of antibiotics generally led to a reduced diversity on plates but yielded different bacteria than other media. In addition, diversity patterns of isolates from agar plates, liquid cultures, and floating filters were significantly different. Almost 89% of all isolates were Alphaproteobacteria; however, members of phyla that are less commonly encountered in cultivation studies, such as Planctomycetes, Verrucomicrobia, and Deltaproteobacteria, were isolated as well. The sponge-associated bacteria were categorized into three different groups. The first group represented OTUs that were also obtained in a clone library from previously analyzed sponge tissue (group 1). Furthermore, we distinguished OTUs that were obtained from sponge tissue (in a previous study) but not from sponge isolates (group 2), and there were also OTUs that were not obtained from sponge tissue but were obtained from sponge isolates (group 3). The 17 OTUs categorized into group 1 represented 10 to 14% of all bacterial OTUs that were present in a large clone library previously generated from Haliclona (gellius) sp. sponge tissue, which is higher than previously reported cultivability scores for sponge-associated bacteria. Six of these 17 OTUs were not obtained from agar plates, which underlines that the use of multiple cultivation methods is worthwhile to increase the diversity of the cultivable microorganisms from sponges.     

REVIEWS

Book reviewed in this article:
Plant Cells . By F. A. L. C lowes and B. E. J uniper .
The History of British Vegetation . By W inifred P ennington . Modern Biology: General Editor J. E. W ebb .
Flora Europaea , Vol. II. Rosaceae to Umbelliferae . Ed. by T. G. T utin , V. H. H eywood , N. A. B urges , D. M. M oore , D. H. V alentine , S. M. W alters and D. A. W ebb .
Desert Wild Flowers . By E dmund J aeger .
British Mosses and Liverworts . By E. V. W atson .
Anatomy of the Monocotyledons , Vol. III. Commelinales—Zingiberales . By P. B. T omlinson . General Editor C. R. M etcalfe .
Biological Science—The Web of Life . Supervising Editor, D. M organ .
Plant Chimeras . By W. N eilson -J ones .
The Potatoes of Argentine, Brazil, Paraguay and Uruguay, a Biosystematic Study . By J. G. H awkes and J. P. H jerting .
Towards an Understanding of the Mechanism of Heredity . By H. L. K. W hitehouse .
Viruses in Plant Hosts; Form, Distribution and Pathologic Effects . By K atherine E sau .
An Introduction to Industrial Mycology . By G eorge S mith .     

Reviews

Plant Lectins. By A. P ustzai .
Techniques for the Rapid Detection of Plant Pathogens . Ed. by J. M. D uncan and L. T orrance .
Ecological Genetics and Air Pollution. Ed. by G. R. T aylor , Jr., L. F. P itelka and M. T. C legg .
Arctic Ecosystems in a Changing Climate. Ed. By F. S. C hapin III, R. L. J efferies , J. F. R eynolds , G. R. S haver , J. S voboda and E. W. C hu .     

REVIEWS

Book reviewed in this article:
Plant and Soil Water Relationships. By P aul J. K ramer .
Vegetable Gums and Resins. By F. N. H owes .
Principles of Human Genetics. By C urt S tern .
Experimental Studies on the Nature of Species. III. Environmental Responses of Climatic Races of Achillea. By J. C lausen , D. D. K eck , W. H. H iesey .
Mountains and Moorlands. By W. H. P earsall .
Spät-und nacheiszeitliche Waldgeschichte Mitteleuropas nördlich der Alpen. Erster Band: Allgemeine Waldgeschichte. By F ranz F irbas .
Biology. An Introduction to Medical and other Studies. By P. D. F. M urray , M.A., D.Sc.
The Fig. By I ra J. C ondit . Foreword by W alter T. S wingle .
American Wild Flowers. By H arold N. M oldenke .     

农业有机废物与城市生活垃圾堆肥高温期微生物种群结构比较

通过PCR、克隆文库方法分析了农业有机废物和城市垃圾堆肥高温期间细菌和真菌种群的多样性.提取堆肥高温期的DNA,PCR扩增,构建各高温期的16S rDNA和18S rDNA克隆文库,结果表明:农业有机废物和城市生活垃圾16S rDNA克隆文库中分别共有18个、21个OTUs,分别属于细菌域的14个、15个不同属,其18S rDNA克隆文库中分别共有8个、9个OTUs,分别属于细菌域的8个、9个不同属,推断农业有机废物堆体的优势菌为Bacillus megaterium、Rhizobium sp.、Phanerochaete chrysosporium、Penicillium sp.同属或同种的菌株;城市生活垃圾堆体的优势菌为Bacillus megaterium、Azospirillum sp.、Phanerochaete chrysosporium同种或同属的菌株.     

共存于厌氧真菌分离培养液中瘤胃甲烷菌的检测及其多样性分析

对分离自山羊瘤胃的真菌分离培养液中甲烷菌进行16SrDNA扩增、DGGE分析、RFLP及测序分析,研究共存于真菌分离培养液中甲烷菌的种类及其多样性。DGGE结果显示:从厌氧真菌分离至第45代,甲烷菌多样性指数由1·32降至0·99,相似性最低为34·7%;第45代至62代,多样性指数由0·99升至1·15,相似性最低为89·2%。RFLP多态性分析69个克隆共得到5个操作分类单元,选择其中6个具有代表性的序列进行测序。序列及系统进化分析表明,属于其中3个操作分类单元的克隆最相似菌都是UnculturedarchaealsymbiontPA202,相似性均为95%,没有与这些克隆相似性较高的已培养甲烷菌;属于另外2个操作分类单元的克隆最相似菌都是Unculturedrumenmethanogen956,相似性均为97%,最相似已知菌为Methanobrevibactersp.NT7,相似性为97%。结果表明,真菌培养液中存在目前尚未分离培养的瘤胃甲烷菌。     

Analysis and comparison of the microbial community structures of two enrichment cultures capable of reductively dechlorinating TCE and cis-DCE

In order to study the effect of different chloroethenes (electron acceptors) on the bacterial composition of dechlorinating communities, two reductive dechlorinating enrichment cultures were developed that were able to reduce trichloroethene (TCE) and cis-1,2-dichloroethene (cis-DCE) to ethene using hydrogen as electron donor, respectively. The inoculum for the cultures was material from a methanogenic fluidized bed reactor (FBR), which was originally seeded with digester sludge and showed a stable capacity for tetrachloroethene (PCE) reduction to ethene for over six years. Molecular methods were used to determine and compare the microbial communities of these two enrichment cultures. A clone library of bacterial 16S rRNA genes was generated for each enrichment. The clones were screened into different groups by restriction fragment length polymorphism (RFLP) analysis using two different four base pair recognition restriction enzymes. A total of 12 sequence types were identified by phylogenetic analysis of nearly complete 16S rDNA sequences ( approximately 1450 bp). The sequences were affiliated with six recognized phyla of the domain Bacteria: Firmicutes (low G+C Gram-positives), Chloroflexi (green non-sulphur bacteria), Actinobacteria (high G+C Gram-positives), Bacteroidetes (Cytophaga-Flexibacter-Bacteroides), Nitrospira and Spirochaetes. The results led to the identification of an organism closely related to Dehalococcoides ethenogenes to be the presumptive dechlorinator in both enrichments. Different electron acceptors affected the bacterial diversity and the community profiles of the two enrichments. Most of the sequences identified in our dechlorinating enrichments shared high similarities with sequences previously obtained from other enriched dechlorinating cultures and chlorinated-compound-contaminated sediments or aquifers, suggesting these bacteria may have direct or indirect roles in reductive dechlorination.     

REVIEWS

Review in this article:
Active Transport and Secretion. Symposium of the Society of Experimental Biology. VIII. Edited by R. B rown add J. F. D anielli .
Protoplasmatologia. Handbuch der Protoplasmaforschung. Hrsg. v. L. V. Heilbrunn u. F. Weber. Band 1/1: Die makromolekulare Chemie und ihre Bedeutung für die Protoplasmaforschung. By H. S taudinger and M. S taudinger .
Flora of Egypt. Vol. III. Angiospermae, part Monocotyledons: Liliaceae—Musaceae. By V ivi T ackholm and M ohammed D rar .
Australian and New Zealand Botany. By J. M c L uckie , M.A., B.Sc. (Glas.), D.Sc. (Syd.) and H. S. M c K ee , B.A., D.Phil. (Oxon).
South African Pollen grains and Spores, Part I. By E. M. van Z inderen B akker .     

REVIEWS

Review in this article:
Protoplasmatologia—Handbuch der Protoplasmaforschung . Band II, CI. The Viscosity of Protoplasm. By L. V. H eilbrunn .
Trace Element Problems in Nature . A Symposium in the Botany Department, University of Cape Town. Edited by D r . K. H. S chütte .
Cell, Organism and Milieu. (Seventeenth Growth Symposium). Edited by D orothea R udnick .
Subcellular Particles . Edited by T. H ayashi .
Plant Growth Substances . By L. J. A udus .
Uredineana. Receuil d': études systematiques et biologiques sur les Urédinées du Globe . Tome 5. By A. L. G uyot .
A Textbook of General Physiology . By H. D avson .
Protoplasmatologia . Vol. Ill AI. Le Chondriome de la Cellule Végétale: Morphologie du Chondriome; by P. D angeard .
Analytical Cytology . Edited by R obert C. M ellors .
Blakeslee: The Genus Datura . By A. G. A very , S. S atina and J. R ietsema .
The Royal Botanic Gardens, Kew, Past and Present . By W. B. T urrill .
Drawings of British Plants . Part XIII. Umbelliferae (2), Araliaceae, Cornaceae. By S tella R oss -C raig .
Physiology of Fungi. By V incent W. C ochrane .
Illustrated Moss Flora of Fennoscandia . Vol. II. Musci. Fasc. III. By E lsa N yholm .
An Outline of General Plant Geography . By W. S zafer .
Protoplasmatologia. Handbuch der Protoplasmaforschung . Ed. L. V. Heilbrunn and F. Weber. Vol. VIII; 9a. Polaritút und Inäquale Teilung des pflanzlichen Protoplasten . By Erwin Bünning, Tübingen.     

Accounting for non-independence of extinction probabilities in the derivation of conservation priorities based on Weitzman’s diversity concept

In the original formulation of Weitzman’s diversity concept, extinction probabilities of different operational taxonomic units (OTUs) are implicitly defined to be independent. In this study, it is shown, that dependencies like concurrence or synergistic relations between OTUs can be accounted for by assuming that joint extinction probabilities differ from the product of the extinction probabilities of the interacting OTUs. A fully analytical treatment is provided for the case of two interacting OTUs, and all cases are illustrated with an example data set. For the case of concurring OTUs, the following general results are derived: (a) concurrence between OTUs always reduces the amount of diversity expected to be conserved in the future; (b) concurrence has a more adverse effect on the more endangered OTUs involved; (c) concurrence can change conservation priorities between OTUs; and (d) with high levels of concurrence, investments in the conservation of some of the concurring OTUs can have a negative effect on the overall diversity conserved. In addition to conservation activities targeted towards some of the OTUs, reduction of the causes of concurrence may be a valid and cost-efficient alternative. For the case of synergistic dependencies between OTUs it was found, that such dependencies always increase the conservation potential of the involved OTUs but may lead to a change of priority ranking in extreme cases. Finally possible extensions and generalisations of the suggested approach are discussed and it is argued, that by adopting these extensions Weitzman’s diversity concept becomes an even more powerful and flexible tool to derive conservation priorities both in an ecological and in a farm animal context.     

Reviews

Ecological Methods with Particular Reference to the Study of Insect Populations. By T. R. E. S outhwood .
Annual Review of Phytopathology , Vol. III. (Ed. J. G. H orsfall .)
Sand and Water Culture Methods used in the Study of Plant Nutrition. (Technical Communication 22, 2nd edition, revised.) By E. J. H ewitt .
The Use of Biological Literature. By R. T. B ottle and H. V. W yatt .