We can find a lot of microorganisms living in snow including psychrophilic snow algae from the order Chlamydomonadales (Chlorophyta). They are adapted to the extreme conditions in this habitat and can cause the phenomenon of coloured snow. The species Chlamydomonas nivalis (Bauer) Wille is the most commonly associated with red snow in alpine and polar regions during summer season worldwide. In the field material, we can find red spherical cells without flagella and any morphological characteristics suitable for species determination. Until now, this species has not been isolated into laboratory culture and its life cycle is unclear. Furthermore it has been shown that red coloured snow can be caused by more species which used to be determined as Chlamydomonas nivalis. The aim of this study was to collect samples of red snow from different parts of Europe, to describe the morphological variability of Chlamydomonas nivalis-like snow algae in relation to region of origin, to try to isolate laboratory strain of this species and to describe its position and distribution by phylogenetic analysis of laboratory strains and field samples. Red snow samples were collected from 30 European localities in Slovenian Alps, Romania, Dolomites, Ötztal, Wallis and Sarntal Alps, High Tauern, Ortler massif, in Norway,...

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The initiation, bloom, and bust of harmful Cyanobacteria and algae blooms (HAB) in lakes are controlled by top-down and bottom-up ecological controls. Excess phosphorous and nitrogen inputs from anthropogenic sources are primary to blame, but eukaryotic grazers may also promote or curb Cyanobacteria dominance. We tracked shifts in bacterial composition, lake chemistry, and eukaryotic grazing community weekly or bi-weekly through spring and summer and modeled the causes of specific Cyanobacterial species blooms and busts across three lakes in Utah, USA, with differing lake trophic states. Regardless of trophic status, all three lakes experienced blooms of varying composition and duration. Aphanizomenon strain MDT14a was the most dominant species in every bloom on Utah Lake, comprising up to 44.16% of the bacterial community. Utah Lake experienced a total of 18 blooms across all sites ranging in duration from one to six weeks. Phormidiaceae sp. (8.5  6.1%) and Microcystis sp. (9.7  4.7%) were the most abundant species in the Deer Creek bloom. Deer creek experienced one bloom at the beginning of fall. Nodularia sp. (9.7  2.1) dominated Great Salt Lake bloom. The Great Salt Lake experienced four separate blooms during the summer months that lasted one to three weeks. Phosphorous concentrations on Utah Lake varied across site and season. Nitrate concentrations on Deer Creek increased over season with a ten-fold increase in concentration. We characterized Cyanobacteria blooms as either bloom communities (growing populations of Cyanobacteria) or as bust communities (declining populations of Cyanobacteria). Using these designations, we modeled the growth and decline of the Cyanobacteria populations across season with top-down and bottom up-controls. Based on generalized least-squared modeling, eukaryotic grazing does not affect relative Cyanobacteria abundances as much as nutrient limitations. Aphanizomenom strain MDT14a was positively correlated with temperature (P < 0.028) and the concentration of K (P = 0.007) and negatively correlated with increases in conductivity (P = 0.0088). Microcystis was positively correlated with increasing levels of SRP (P < 0.001) and negatively correlated with higher Ca concentrations (P = 0.008) and PP (P = 0.008). Busts of Microcystis were related to decreases in nitrate (P = 0.06) and lower total lake depths (P = 0.03). Phormidiaceae sp. relative abundance was negatively correlated with higher levels of TDN (P = 0.01-0.001) and Mg (P = 0.01) and positively correlated with higher S concentrations (P = 0.007). Our findings suggest that micronutrients and more bioavailable forms of P may potentially allow Cyanobacteria to break dormancy and proliferate HAB communities.

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This social history of science policy development emphasizes the impact on the agricultural community of federal policies regarding release of recombinant DNA (rDNA) organisms into the environment. The history also demonstrates that the U.S. Coordinated Framework for Biotechnology Regulation (1986) is based principally in political criteria, not solidly based in science as its proponents claimed. The power struggle among policy negotiators with incompatible belief systems resulted in a political correction of biotechnology. I also demonstrate that episodes in the rDNA controversy occur in repetitive and periodic patterns. During the 1980s, the first rDNA microbial pesticide, Ice-Minus, struggled through a policy gauntlet of federal agency approval processes, a Congressional hearing, and many legal actions before it was finally released into the environment. At the height of the controversy (1984-1986), the Reagan Administration would admit no new laws or regulations to slow the development of technologies or hinder American international competitiveness. At the same time, Jeremy Rifkin, a radical activist representing a green world view, used the controversy to agitate for social and economic reform. Meanwhile, a group of Congressional aides who called themselves the "Cloneheads" used the debate to fight for more public participation in the science policy-making process. Conflicting perspectives regarding biotechnology originated, not in level of understanding of the science involved, but in personal perspectives that were outwardly expressed as political group affiliations. The direction of federal biotechnology policy was influenced most successfully by politically best-positioned individuals (what I call a "hierarchy effect") who based decisions on how biotechnology harmonized with their pre-existing beliefs. The success of their actions also depended on timing. Historical events during the rDNA controversy followed the same periodic pattern--gestation, threshold, crisis/conflict, and quasi-quiescence--through two consecutive eras--the Containment Era (1970s) and the Release Era (1980s). These periods are modeled after Fletcher's stages through which ethical issues evolve (1990). However, an agricultural perspective on the debate reveals that such stages also occur in finer detail on repeating, overlapping, and multi-level scales. Knowledge of this periodicity may be useful in predicting features of future episodes of the rDNA controversy. / Ph. D.

Chlorbenzole sind aufgrund ihrer weiten Verbreitung in Industrie und Landwirtschaft und ihrer geringen chemischen Reaktivität sowie guten Fettlöslichkeit ubiquitäre Umweltkontaminanten, die sich in der Nahrungskette anreichern. Eine natürliche, mikrobielle Dechlorierung dieser Verbindungen ist von besonderem Interesse, da die Toxizität chlorierter Benzole mit der Anzahl der Chlorsubstituenten steigt. Im Gegensatz zu Mono- und Dichlorbenzol, die durch aerobe Laborreinkulturen dechlorierbar sind, werden höher chlorierte Benzole bevorzugt unter anaeroben Bedingungen in Mischkulturen mit unbekannter Spezieszusammensetzung dechloriert. Bioreaktoren, in denen solche undefinierten Mischkulturen kontinuierlich kultiviert werden, sind eine vielversprechende Technik bei der Behandlung Chlorbenzol-kontaminierter Abwässer. Aufgrund der unbekannten Zusammensetzung der Population muß die mikrobielle Aktivität jedoch als "black box" betrachtet werden, was eine direkte Steuerung und Optimierung solcher Bioreaktoren erschwert. Zur Bestimmung der mikrobiellen Diversität einer in ihrer Zusammensetzung unbekannten, Trichlorbenzol(TCB)-dechlorierenden Bioreaktorkultur wurde aufgrund der bekannten Limitierungen kulturabhängiger Methoden die kulturunabhängige, vergleichende Sequenzanalyse direkt amplifizierter und klonierter 16S-rDNA gewählt. Die durch eine neuentwickelte Hybridisierungsmethode wesentlich vereinfachte Analyse der 16S-rDNA-Genbibliotheken erlaubte eine phylogenetische Klassifizierung der in der TCB-dechlorierenden Mischkultur abundanten Mikroorganismen (Bakterien und Archaea) und zeigte das Auftreten von 51 bakteriellen 16S-rDNA-Klonfamilien, mit einem weiten phylogenetischen Spektrum und teilweise enge Verwandtschaften zu anaerob dechlorierenden Dehalobacter spp. oder zu unkultivierten Bakterien eines vergleichbaren Biotops. Dieser Diversität stand eine dominierende Methanosaeta-ähnliche Klonfamilie innerhalb der Archaea gegenüber. Die aus der phylogenetischen Klassifizierung abgeleiteten metabolischen Eigenschaften einiger Bakterien und Archaea der TCB-dechlorierenden Mischkultur konnten durch gezielte Anreicherung und in-vitro Kultivierung bzw. die kulturunabhängige Sequenzanalyse funktioneller Gene bestätigt werden. Die dargestellten Ergebnisse lassen eine spezifische Zusammensetzung der TCB-dechlorierenden Mischkultur vermuten und geben Hinweise auf Indikatororganismen, die ein Monitoring und eine damit verbundene Effizienzsteigerung der anaeroben TCB-Dechlorierung im Bioreaktor ermöglichen könnten. / Due to their widespread application in industry and agriculture and their chemical stability chlorobenzenes (CB) are ubiquitous pollutants in soil, sediments, and aquifers. Since toxicity of CBs increases with the number of chlorine substituents, microbial dechlorination of CBs is of major interest. In contrast to di- and monochlorobenzenes (DCB and MCB) higher chlorinated benzenes are more resistant to aerobic dechlorination. However, for these compounds reductive dechlorination by different anaerobic microbial communities is well known. Bioreactors inoculated with complex dechlorinating anaerobic microbiota seem to be promising technologies for bioremediation of CB-contaminated aquifers. Several studies showed the efficiency of such bioreactors in treating chloroaromatic contaminated wastewaters. However, due to the unknown species diversity microbial activity had to be treated as a "black box" and direct optimization was hampered. To determine the microbial diversity of an anaerobic consortium in a fluidized bed reactor used for dechlorination of trichlorobenzene (TCB) I employed comparative sequence analysis of 16S rRNA genes after direct PCR-amplification and cloning from community DNA since culture-based methods have shown to be strongly biased. The application of a new hybridization based screening approach for bacterial 16S rDNA clone libraries drastically simplified the analysis and allowed the phylogenetic classification of the abundant bacteria and archaea. A total of 51 bacterial 16S rDNA clone families were found, which showed a wide distribution among the main bacterial phyla. Several bacterial 16S rDNA clone families were closely related to anaerobic, dechlorinating Dehalobacter spp. and to yet-uncultured bacteria of a similar habitat. In contrast to the high bacterial diversity archaeal 16S rDNA clone libraries were clearly dominated by a Methanosaeta concilii-like clone family. Some yet-uncultured bacteria and archaea of the TCB-dechlorinating consortium were sufficiently closely related to studied organsims that reasonable physiological hypotheses could be formulated. These hypotheses were confirmed by either cultivation of the respective organism or by culture-independent sequence analysis of specific functional genes. The results suggest a specific community structure of the TCB-dechlorinating consortium and give evidence for indiator organisms. Moleculargenetic monitoring of these indicator organisms might allow the optimization of the continous TCB-dechlorination in the fluidized bed reactor.

Made available in DSpace on 2015-03-26T13:51:43Z (GMT). No. of bitstreams: 1 texto completo.pdf: 248917 bytes, checksum: 84bc05c660ca867229222e12c0fa08ec (MD5) Previous issue date: 2003-03-31 / Fundação de Amparo a Pesquisa do Estado de Minas Gerais / Phylogenetic analysis of Lactobacillus UFV H2b20 based on the 16S rDNA sequence positions this isolate among the Lactobacillus delbrueckii subspecies. It confirms the classification by DNA/DNA hybridization. Comparisons of the substituted bases along the 16S rDNA sequence show that most are probably caused by 5-methylcytosine deamination that results in GC/AT transition. Presence of polymorphic copies of the 16S rRNA gene in Lactobacillus delbrueckii UFV H2b20 were confirmed by PCR-DGGE. Five distinct bands of rDNA amplified sequences confirm at least five different copies of the gene. The intensity of one of the bands allows the inference of two identical copies. The DGGE profile of this isolate was different from those of other L. delbrueckii strains, providing a mean to distinguish it in mixed cultures. Analysis of the nucleotide sequences along the rDNA region revealed the presence of putative insertion sequences. One of them, 915 bp long, was characterized and denominated ISLdH2b20. It presents a single ORF that encodes a putative transposase with some homology to the one of ISL5 of L. delbrueckii subsp. bulgaricus. All the characteristic elements of an IS were located as well as the putative regulatory sequences of the transposase. / O estudo de filogenia baseado em seqüências de rDNA de Lactobacillus UFV H2b20 mostrou que esse microrganismo foi agrupado entre as subespécies de Lactobacillus delbrueckii. Este resultado confirma a nova classificação do isolado, baseada na hibridização DNA-DNA. A análise das substituições de bases mostra que a maioria é causada pela desaminação da 5-metilcitosina, o que causa uma transição GC/AT. A técnica PCR-DGGE de L. delbrueckii UFV H2b20 demonstrou resultados satisfatórios na análise de polimorfismos do operon rrn em microrganismos isolados. Foram encontradas cinco bandas diferentes para o L. delbrueckii UFV H2b20, confirmando a presença de cinco cópias distintas do rDNA 16S nesse microrganismo e possivelmente seis no total. O isolado L. delbrueckii UFV H2b20 apresentou um perfil de bandas diferente das outras linhagens de Lactobacillus, o que poderá ser utilizado para diferenciá-lo dos demais, uma vez que outros métodos baseados em PCR, como RAPD, mostram-se pouco discriminatórios. A análise das seqüências da região do rDNA 16S revelou uma seqüência de inserção putativa, de 915 pb, que foi caracterizada e denominada ISLdH2b20. Ela apresenta uma única ORF e codifica uma transposase putativa que apresenta homologia com a transposase de ISL5. Todos os elementos característicos de uma IS foram identificados bem como as seqüências regulatórias putativas da transposase.

La mer Méditerranée est une des zones océaniques les plus oligotrophes de la planète pour laquelle la fixation biologique d’azote a été proposée comme jouant un rôle important dans les flux de carbone et d’azote. Comme les organismes planctoniques potentiellement responsables de cette activité n’y ont jamais été étudiés, les objectifs de cette thèse étaient de caractériser la diversité (abondance et richesse spécifique), la distribution spatio-temporelle, et les facteurs de contrôle des diazotrophes, en s’axant particulièrement sur les cyanobactéries unicellulaires (UCYN). Pour cela, des hybridations in situ (TSAFISH)sur différentes fractions de taille en combinaison avec des comptages microscopiques et des analyses phylogénétiques sur les gènes 16S ADNr et nifH ont été réalisées, à la fois lors d’un cycle saisonnier à la station côtière SOMLIT de Marseille, et lors du transect Méditerranéen BOUM. Alors que les cyanobactéries filamenteuses Richelia intra cellularis et Trichodesmium sp. ont été détectées seulement ponctuellement dans le temps et dans l’espace, en très faible concentration (0.02 filament.ml-1), la communauté des cyanobactéries diazotrophes était dominée à 99,9% par du picoplancton hybridé avec la sonde Nitro821, spécifique aux UCYN. Ces cellules, de petite (0.7-1.5 μm) ou de grande (2.5-3.2 μm)taille, ont été retrouvées en faible concentration (1-6 cellule.ml-1) dans toute la Méditerranée et tout au long de l’année, à l’exception de l’été 2006 lorsque la concentration des petites cellules a atteint 5300 cellule.ml-1 au cours d’un évènement exceptionnel de pollution atmosphérique urbaine. Des efflorescences similaire sont été observées suite à des enrichissements en poussières sahariennes à une station côtière en Corse et au large au centre de la Méditerranée, simultanément avec des augmentations de l’activité de fixation d’azote.L’affiliation des petites cellules aux UCYN-A a été confirmée par phylogénie dans les eaux du large de la Méditerranée occidentale. Les librairies de clones nifH de la fraction de taille picoplanctonique étaient dominées par des séquences d’α-protéobactéries appartenant à un nouveau groupe marin de Bradyrhizobium. D’autres groupes de rhizobia et des γ-protéobactéries ont été détectés ponctuellement.Alors que l’absence de Trichodesmium sp. au large pourrait être liée aux faibles concentrations en phosphate et en fer, les facteurs inhibant le développement des UCYN-B et –C restent inconnus. Lesdiazotrophes picoplanctoniques (Bradyrhizobium, rhizobia, UCYN-A) pourraient avoir développé des stratégies spécifiques, telles que des associations avec des eucaryotes ou des particules inertes, et/ou la capacité de photosynthèse, pour acquérir le carbone nécessaire au soutien du processus de la diazotrophie / The Mediterranean Sea is one of the most oligotrophic marine areas on earth where nitrogenfixation has been formally believed to play an important role in carbon and nitrogen fluxes. Although thisview is under debate, the diazotrophs responsible for this activity have still not been investigated. The aimsof this PhD were to characterise the diversity (abundance and species richness) and the spatio-temporaldistribution of diazotrophs, as well as factors controlling their development, with a particular focus onunicellular cyanobacteria. A combination of microscopic counts with size fractionated in situ hybridization(TSA-FISH), and 16S rDNA and nifH phylogenies were done, either over a year and a half seasonal cycle atthe coastal SOMLIT station off Marseilles, and across the entire Mediterranean Sea during the BOUMtransect. Low concentrations of diazotrophic cyanobacteria were detected and this community wasdominated at 99.9% by picoplankton hybridized with Nitro821 probe, specific for unicellular diazotrophiccyanobacteria (UCYN). Among filamentous cyanobacteria, only 0.02 filament ml-1 of Richeliaintracellularis and Trichodesmium sp. were detected sporadically in time and space. Small (0.7-1.5 μm) andlarge (2.5-3.2 μm) Nitro821-targeted cells were recovered in low concentrations (1-6 cell ml-1) across theentire Mediterranean Sea and all the year long, except over a month period in summer 2006 whenconcentrations of small cells reached 5300 cell ml-1, during an exceptionally high urban pollution event.Similar blooms of small and large cells were reported after Saharan dust inputs off Corsica and at open Sea,simultaneously with increases in N2 fixation rates. The affiliation of the small Nitro821-targeted cells toUCYN-A was confirmed by 16S and nifH phylogenies offshore in the western Mediterranean Sea. Rhizobiasequences, including the ones of a new marine group of Bradyrhizobium, were dominating nifH clonelibraries from picoplanktonic size fractions. A few sequences of γ-proteobacteria were also detected incentral Mediterranean Sea. While low phosphate and iron concentrations could explain the absence ofTrichodesmium sp. offshore, the factors that prevent the development of UCYN-B and C remain unknown.It is proposed that the dominating Mediterranean picoplankters (Bradyrhizobium, rhizobia, UCYN-A)probably developed specific strategies, such as associations with protists or particles, and/or photosyntheticactivity, to acquire carbon for sustaining diazotrophy.

Depuis l’apparition de la vie sur terre, les pressions de sélection liées aux interactions biotiques et abiotiques ont généré une forte diversité des formes de vie. Ainsi, chaque espèce eucaryote coévolue avec sa communauté microbienne associée. Dans le cas des plantes, la diversité génétique se traduit au niveau de multiples traits phénotypiques (exsudation de substrats carbonés, architecture racinaire, densité et aération du sol, acidification, etc.) susceptibles d’influer sur les interactions avec les populations microbiennes du sol, et donc sur la composition et le fonctionnement de la communauté microbienne rhizosphérique. Notre hypothèse est que les différences entre communautés bactériennes rhizosphériques sont proportionnelles aux distances évolutives entre partenaires végétaux. L’objectif de cette thèse était donc de déterminer l’importance, dans le cas des Poacées et notamment du maïs, de l’histoire évolutive de la plante dans la capacité de sélection des communautés bactériennes de la rhizosphère. Les analyses faites à l’aide d’une puce à ADN taxonomique 16S indiquent que la composition de la communauté rhizobactérienne dépend du groupe génétique de maïs mais n’est pas liée aux marqueurs microsatellites de diversité du maïs. Par contre, à l’échelle des Poacées, une corrélation a été trouvée entre la phylogénie végétale et la composition de la communauté bactérienne (voire la prévalence de taxons bactériens particuliers). Cette corrélation n’était pas significative quand l’étude était limitée à l’effectif, le niveau de transcription de nifH ou la diversité du groupe fonctionnel des bactéries fixatrices d’azote. En conclusion, l’histoire évolutive du partenaire végétal à l’échelle des Poacées (mais pas à celle du maïs) est un facteur conditionnant les interactions avec les groupes bactériens taxonomiques (mais pas nécessairement fonctionnels) de la rhizosphère / Since the emergence of life on earth, the selection pressures related to biotic and abiotic interactions generated a high diversity of life forms. Thus, each eukaryotic species co-evolved with its associated microbial community. In the case of plants, genetic diversity is reflected in many phenotypic traits (exudation of carbon substrates, root architecture, soil density, aeration, acidification, etc.), and may influence interactions with soil microbial populations and hence the composition and functioning of the rhizosphere microbial community. Our hypothesis is that the differences between rhizosphere bacterial communities are proportional to evolutionary distances between plants partners. The objective of this thesis was to determine the importance, in the case of Poaceae and in particular of maize, of the evolutionary history of plant in the selection of bacterial communities in the rhizosphere. Analyses performed using a 16S taxonomic microarray indicated that the composition of the rhizobacterial community depends on the genetic group of maize but is not linked to microsatellite diversity of maize. Conversely, across the Poaceae, a correlation was found between plant phylogeny and the composition of the bacterial community (and the prevalence of specific bacterial taxa). This correlation was not significant when the study was limited to the size, the level of transcription or nifH diversity of the functional group of nitrogen-fixing bacteria. In conclusion, the evolutionary history of the plant partner across the Poaceae (but not maize) is a factor conditioning interactions with bacterial taxonomic groups (but not necessarily functional groups) in the rhizosphere