The early Pliocene age Gray Fossil Site (GFS) was relatively recently found, with much still to be discovered and examined, and represents one of only a few sites of its age in eastern North America. It has been noted that the diverse faunal remains found at GFS are unique compared to what have been found at other fossil sites in North America from the same time period. Studying mammalian community structures at fossil sites can provide an abundance of information about the past environment and species adaptations to it, such as niche occupation of species, resource partitioning, and interactions between organisms and their environment. The main questions being asked in this study are: 1) what is the mammal community structure like at GFS?, and 2) how does the community structure at GFS compare to other contemporaneous sites? While studies of the fauna and flora have helped us to understand the ecosystem at GFS, detailed study of the mammal community will help us better understand this unique site. It is expected that the community structure and niche occupation of the mammals at GFS will be different from other contemporaneous sites, dominated by mammals adapted for life in the warm oak, hickory forest present at the site. A taxon-free approach to analysis will be used so that environments not sharing the same taxa or of different ages can be compared to one another. Each mammal species from GFS (48 total) and five well-known contemporaneous sites are categorized by body size, locomotor mode, and dietary preference to characterize the niches occupied by each species. Categorizations for each species will come from published works and measurements / ecomorphological analysis of specimens. Preliminary results show that GFS is different from other sites in that there are more brachyodont and tree dwelling/climbing adapted species present, and there are fewer running species present. The initial findings are likely associated with the site being a closed, forested ecosystem, compared to the more open environments of the other sites. Moving forward, descriptions pertaining to specifically how GFS and its mammal community structure compare to the niche occupation of similar species at other sites will be discussed. This project will also examine what more this analysis can reveal about the ecosystem at GFS, particularly how the herbivorous mammals can provide greater insight into what environmental conditions were like, including what vegetation was predominant at GFS.

With an increase in demand on deep-sea resources comes a need for appropriate and effective management of this ecosystem. The establishment of a representative network of deep-sea Marine Protected Areas offers one tool with which to address the conservation needs of the deep sea. While a number of deep-sea habitats have been identified as vulnerable to anthropogenic activities (e.g. cold-water coral reefs and sponge aggregations), poor knowledge of the distribution of these habitats hinders conservation efforts and network planning, and thus we need habitat maps. With improvements in acoustic data resolution acquired from the deep sea, and the ability to cover large areas rapidly, the use of acoustic techniques in mapping biological habitats is growing. Multibeam bathymetry and its derived terrain variables can potentially provide important information that can aid in the delineation and characterisation of biological communities. A necessary prelude to mapping is therefore the definition of biological assemblages for use as mapping units. Two megahabitat features (seamount and submarine canyons) were sampled using acoustic and ground-truthing to characterise and map the distribution of benthic assemblages. Species were identified as distinct morpho-types and catalogued, and still images quantitatively analysed. Standard multivariate community analysis was undertaken to define distinct faunal assemblage that may act as mapping units. Those clusters identified by the SIMPROF routine were taken against a set of criteria to reject/accept as robust assemblages that may be used as mapping units. Twenty two benthic assemblages or biotopes were defined from multivariate analysis of quantitative species data, 11 from the SW Approaches and 11 from Anton Dohrn Seamount, and a further one from video observations (SW Approaches). Taken against current definitions, 11 of these were considered as Vulnerable Marine Ecosystems (VME). Diversity was measured to compliment the comprehensive description of biotopes. The use of multivariate diversity indices proved better for comparing diversity of biotopes as it captures a more than one aspect of diversity of the community. Two biotopes were common to both megahabitat features, cold-water coral reef habitats, and those from Anton Dohrn Seamount were more diverse than from the SW Approaches. Modelling techniques were employed to test the relationship between biotopes and environmental and geophysical parameters, which may be used as surrogates to map VME. Generalised Additive Models of Vulnerable Marine Ecosystems revealed multibeam bathymetry and its derived parameters to be significant surrogate for mapping the distribution of some assemblages, particularly those that appear to be influenced by current regime; whilst not so well for those whose distribution is not so strongly current driven e.g. soft sediment communities. In terms of deep-sea mapping, the use of multibeam can prove a useful mapping tool if the resolution of the data is at an appropriate scale that will identify meso-scale geomorphological features, such as cliff-top mounds, that may act as proxies for occurrence of biotopes, but this relationship is still unclear. Surrogates were used to map VME across the seamount and submarine canyons, and full coverage maps were produced for all biotopes occurring on these megahabitat features.

The content in my graduation theses is characteristic of villages and poeple in outbuildings , which disappeared by the development JETE. I wrote about trend in number of houses and poeple in the time 1869 {--} 2001 in comparision with Southbohemia region, district around Č. Budějovice, city Týn n. Vltavou and village Temelín, which is nearly JETE. People compare target the age, nationality and economy power in the country around JETE. I wanted to desribe structure of settled outbuildings. The trend of the country target the farmland, status quo around JETE, i.c. reconstruction of castle Býšov in distillery, characteristic park of castle, valuation of landscape and coefficient ecological stability.

This study examined aspects of local reproductive variation in the flowering dogwood (Cornus florida L.) coincident with recent differences in primary canopy architecture. The dogwood trees in this study were impacted by a hurricane that created numerous treefall gaps, which created fine scale heterogeneity in the primary canopy. Fine scale disturbances in a forest can result in changes for multiple members of the forest community, including the reproductive responses of the trees and interspecific pollination mutualisms. Previously determined differences in offspring genetic structure suggested that pollen movement among genetically unstructured maternal individuals was significantly impacted between open, or disturbed, canopy and closed, or undisturbed, types. To further understand mechanisms by which this nonrandom mating occurred, this study examined both intrinsic and extrinsic reproductive factors for C. florida. The first chapter examines several parameters of the reproductive phenology of sample groups of dogwood between the canopy types as intrinsic factors. The parameters observed included initiation, course and termination of flowering, seed set and seed germination ratios. The results showed some significant differences between the sampling areas in flowering intensity, although the majority of the variation was between maternal individuals throughout the entire sample. The second chapter examines insect community composition across areas of differential canopy disturbance. Community analysis showed that the open canopy contained a larger and more diverse assemblage of insects than the closed canopy or the field, which represented an area of complete canopy removal. This shift in insect community composition may have created functional differences in the local pattern of pollen flow by altering the functional composition of local potential pollinator assemblages. The major finding of this study was that the impact of intermediate natural disturbance on the mating systems of understory plant populations may be more indirect than direct. In conclusion, the observed differences in insect community structure within these habitat types in this study were consistent with predetermined patterns of pollen flow; this structure can explain some of the previously observed genetic structure within locally proximate understory tree populations.

Nitrogen is known to be a limiting factor in polluted environments, however many studies overlook the potential role for nitrogen to significantly influence the removal rate and efficiency with which microorganisms can degrade aromatic hydrocarbons. In this study, inoculated and uninoculated aerated soil microcosms containing different s-triazines were examined for their ability to efficiently and rapidly treat contaminated soils containing naphthalene, nitrobenzene, and toluene (NNT), via a microbial consortium consisting of Pseudomonas, Rhodococcus, and Aeromonas. After an experimental period of 14 days, greater than 90% degradation of NNT supplemented with different s-triazines, at concentrations of 1000-3000 ppm was observed. A difference in the degradation of NNT was seen in inoculated box reactors supplemented with cyanuric acid, melamine, and atrazine in comparison to uninoculated box reactors. Combined usage of 16s rDNA and 16s rRNA analysis was then applied to study the bacterial communities, and determine the abundance and survival of inoculated strains within box reactors contaminated with NNT. The bacterial diversity within clone libraries obtained illustrated a dominance of proteobacteria and gram positive bacteria. Analysis from clone libraries also showed that inoculated strains did survive within each condition, but were not the most predominant members present in the communities. This research shows that significant removal of NNT can be achieved in two weeks with the supplementation of one of the s-triazines. However, differences in degradation and the microbial populations present within contaminated communities will be seen depending on which nitrogen sources are used and whether or not environments are bioaugmented or not.

Contaminated aquatic sediments pose health risks to fish, wildlife, and humans and can limit recreational and economic uses of surface waters. Technical and cost effective in situ approaches for sediment management and remediation have been identified as a research need. Subaqueous in situ capping is a promising remedial approach; however, little is known regarding its impact on underlying sedimentary processes and the feasibility of bioaugmented caps at sites subject to contaminated groundwater seepage. This work specifically addresses (1) the impact of capping on biogeochemical processes at the sediment-water interface, (2) the ability and degree to which indigenous sediment microorganisms colonize an overlying cap, (3) the effect of advective flow direction on redox conditions within a cap, (4) natural contaminant bioattenuation processes within capped sediment, and (5) limitations toward a functional bioreactive in situ cap. Laboratory-scale experiments with capped sediment columns demonstrated that emplacement of a sand-based in situ cap induced an upward, vertical shift of terminal electron accepting processes into the overlying cap while simultaneously conserving redox stratification. Upflow conditions simulating a groundwater seep compressed anaerobic processes towards the cap-water interface. Microorganisms indigenous to the underlying sediment colonized cap material and spatial population differences generally reflected redox stratification. Downflow of oxic surface water through the cap, simulating tidally-induced recharge, created fully oxic conditions within the cap, demonstrating that flow direction strongly contributes to redox conditions. Experiments simulating capped sediment subject to contaminated groundwater seepage revealed a reduction of natural bioattenuation processes with time, stemming from the elimination of labile organic matter deposition to the sediment and a subsequent lack of electron donor. Thus, parent contaminants within groundwater seeps will be subject to minimal biotransformations within the sediment before entering a reducing cap. A bioreactive cap, inoculated with microorganisms capable of reductive dehalogenation, was established to reductively dechlorinate tetrachloroethene present in the groundwater; however electron donor amendments to sediment effluent were required to achieve complete dechlorination of tetrachloroethene to non-toxic ethene. Results from this work improve understanding of biogeochemical and bioattenuation processes within capped aquatic sediments and should aid in the development of active capping technologies.

In chapter 1 I describe the population dynamics of an understudied species of gulper shark, Centrophorus uyato (common name, the Little Gulper), found in the Northern Gulf of Mexico. Sharks in the family Centrophoridae are mid-sized, demersal fish, with seven species identified in North American waters. These deepwater species can be difficult to study due to the extreme depths at which they occur. During four longlining cruises from 2012-2014, 593 sharks were landed, predominantly in the Mississippi Canyon off the Louisiana coast. Mean depth of capture was 290 m. Supplementing these data are catch records for C. uyato from a second series of cruises east of these stations. These data suggest the Little Gulper displays sexual dimorphism in its population size structure, similar to other elasmobranchs. Length frequency distributions, sex ratios by length, and length-weight curves were similar for both catch series, however females comprised a greater fraction of the catch from Mississippi Canyon stations compared to those further east. The high incidence of females in the region may indicate the use of Mississippi Canyon as a nursery for C. uyato, and the species' tendency to sexually segregate at some point in its life history. A high number of large, pregnant females, along with observations of recently delivered animals, was noted at the Mississippi Canyon sites. Little Gulper sex ratios by length exhibited a characteristic "notch", indicative of a low female-to-male ratio at intermediate lengths, then a reversal to female dominance at the largest sizes captured. I modeled this curve under various assumptions of growth, mortality, and longevity for the species. The combination of the von Bertalanffy growth parameter, K, and the total instantaneous mortality rate, Z, that best fit the data suggests a population of slow growing, long-lived animals with a differentially higher growth rate for males. In chapter 2, I examine species associations among elasmobranchs in the Northern Gulf of Mexico in relation to depth of capture. From 2011-2014, 3609 sharks, comprising 31 species in 10 families, were landed on five long-lining cruises throughout the Northern Gulf and along the West Florida Shelf. Stations were grouped a priori according to depth into one of three categories, based on pre-existing divisions in the station data: 0-73 m (shallow), 110-146 m (intermediate), and 183 m or greater (deep). Using a non-parametric multivariate analysis of variance (NP-MANOVA) and a pairwise test found I found significant differences in species composition among all three depth groupings. Gulf Smoothhound (Mustelus sinusmexicanus) and Atlantic Sharpnose (Rhizoprionodon terraenovae) contributed most to the differences, and also accounted for over 65% of the total catch. A canonical analysis of principal coordinates (CAP) showed mid-water species such as the Scalloped Hammerhead (Sphyrna lewini) to be more indicative of intermediate depth stations, while the Little Gulper (Centrophorus uyato) and Shortspine Dogfish (Squalus mitsukurii) distinguished deepwater sites. A "leave-one-out cross-validation" (LOO-CV) procedure correctly re-classified 78% of the samples back into their a priori groupings; catches from the shallow and deep groups were never confused for one another, although there was moderate confusion with classifying catches from intermediate stations. Taking into consideration the potentially large movements undertaken by many shark species on both small and large time scales, as well as additional biological and physical drivers for range and location, it is unsurprising that these animals would be present in two or more depth groups, and thus more frequently misclassified. I also performed a second analysis grouping the stations according to their location on the West Florida Shelf, or in the eastern or western quadrants of the Northern Gulf of Mexico. Again I found significant differences among the species composition of all three station groups, but the LOO-CV overall reclassification success rate was lower than when the stations were grouped according to depth. There was also a smaller interval between this reclassification success rate and one performed using random group allocation. Both of these chapters provide data on previously poorly understood species and phenomena in the Northern Gulf of Mexico. These data are important in defining the potential susceptibility of shark species and communities to future threats, including by-catch in fisheries and contamination events such as shallow and deepwater oil spills.

Oenococcus oeni est une espèce de bactérie lactique adaptée à l'environnement hostile de la fermentation du vin. Elle montre un degré de spécialisation remarquable face au stress provoqué par le faible pH et la forte teneur en éthanol, ce qui lui permet de proliférer là où la plupart des bactéries ne survivent pas. Cette bactérie est très importante dans la production de vin, car elle réalise la fermentation malolactique, qui se produit après la fermentation alcoolique, et au cours de laquelle l'acide malique est métabolisé en acide lactique et où le vin est désacidifié. L'espèce accumule des mutations plus vite que les autres espèces de bactéries lactiques, ce qui a probablement accéléré le processus de domestication. Son degré de spécialisation a été démontré par la présence de populations spécifiques adaptées aux vins rouges ou aux vins blancs dans la même région. Dans cette étude, nous avons utilisé des approches de séquençage haut débit et de génomique pour élucider la diversité des souches d’O. oeni, identifier leurs caractéristiques génomiques et mesurer leur dispersion dans différents environnements ainsi que leur dynamique au cours des fermentations. En raison de son importance pour la vinification, plusieurs centaines de souches ont été isolées et séquencées. Dans ce travail, nous avons augmenté la collection de génomes en séquençant des souches de cidre et de kombucha et en effectuant des analyses phylogénétiques afin de clarifier la structure de la population de l'espèce. En calculant un pangénome à l'échelle de l'espèce, nous avons effectué une analyse génomique comparative afin d'explorer des gènes spécifiques à une ou plusieurs sous-populations. Avec le séquençage de nouvelle génération, nous avons produit des génomes entièrement circularisés à partir des principales sous-populations et analysé leurs arrangements génomiques. Ces nouveaux génomes ont été annotés avec de nouveaux pipelines automatiques et une curation manuelle pour la première fois depuis la publication du génome de référence PSU-1. L’évolution des communautés bactériennes au cours de la fermentation, du moût de raisin au vin fini, a été examinée par le séquençage de fragments 16S dans quatre exploitations du bordelais. À l’aide d’amorces universelles et spécifiques, nous avons comparé la biodiversité des espèces dans des vins issus d’agriculture biologique ou conventionnelle. De plus, en se basant sur les groupes phylogénétiques de souches d’O. oeni nouvellement définis, nous avons développé une méthode de qPCR pour analyser la dispersion des groupes de souches d’O. oeni et leur dynamique au cours des fermentations. Cette nouvelle méthode a également été utilisée pour analyser la diversité des souches d’O. oeni dans les vins de base de Cognac et au cours de la production de cidre, deux produits qui se distinguent des productions de vins traditionnels par la non-utilisation de sulfites. Les deux autres espèces du genre Oenococcus, O. kitaharae et O. alcoholitolerans, se retrouvent également dans les environnements de boissons fermentées. O. kitaharae ne possède pas de gène malolactique fonctionnel, mais O. alcoholitolerans, découvert plus récemment, serait capable de réaliser la réaction malolactique. Nous l’avons caractérisée, ainsi que sa tolérance aux facteurs de stress de l'environnement vin. Constatant qu'elle était incapable de survivre dans le vin, nous avons produit un génome entièrement circularisé d'O. alcoholitolerans et effectué une analyse de génomique comparative afin d'identifier les gènes d'O. oeni lui permettant de tolérer le pH et l'éthanol, ce qui manque à O. alcoholitolerans et à O. kitaharae. En conclusion, nous avons utilisé les nouvelles technologies de séquençage de nouvelle génération pour produire des génomes de haute qualité et effectuer des analyses comparatives approfondies à l’échelle de l’espèce qui nous ont permis d’identifier des gènes susceptibles d’expliquer l’adaptation d’O. oeni à l’environnement. / Oenococcus oeni is a lactic acid bacteria species adapted to the inhospitable environment of fermenting wine, where it shows a remarkable degree of specialization to the stress of low pH and high ethanol that allows it to proliferate where most bacteria fail to survive. The bacteria is supremely important in wine production, because it carries out malolactic fermentation, a process that occurs after alcoholic fermentation, where malic acid is metabolised into lactic acid and the pH of the wine is raised. The species has only a small genome and accumulates mutations several orders of magnitude faster than other lactic acid bacteria due to a loss of DNA mismatch repair genes. This has likely sped up the process of domestication to wine. The degree of specialization has been demonstrated by finding specific populations adapted to red or white wines in the same region. In this study, we used high throughput sequencing and genomics approaches to elucidate the diversity of O. oeni strains, to identify their genomic characteristics and measure their dispersion in different environments as well as their dynamics during fermentation. Because of its importance to wine-making, several hundred strains have been isolated and sequenced. In this work, we have expanded upon the collection of genomes by sequencing strains from cider and kombucha and performing phylogenetic analyses to clarify the population structure of the species. By calculating a species-wide pangenome, we performed comparative genomics to explore gene clusters that were specific to one or more sub-populations. With next generation sequencing, we produced fully circularized genomes from the major sub-populations and analysed their genomic arrangements. These new genomes were annotated with new, automatic pipelines and manual curation for the first time since the publication of the reference genome PSU-1. The evolution of bacterial communities over the course of fermentation, from grape must to finished wine, was examined with 16S amplicon sequencing in four Bordeaux wineries. Using a universal and a specific primer-set, we compared the biodiversity in wines resulting from organic or conventional farming practices. In addition, with the newly defined phylogenetic groups, we developed a qPCR experiment to detail the composition of O. oeni in the fermentations and cemented the dispersal of even rarely isolated strain sub-populations in grape must. This new method was also used to analyse the diversity of O. oeni strains in the base wines of Cognac and during the production of cider, two products that are distinguished from traditional wine production by not using sulfite. The two other species in the Oenococcus genus, kitaharae and alcoholitolerans, are also found in the environments of fermenting beverages. O. kitaharae does not have a functional malolactic gene, but the more recently discovered O. alcoholitolerans was thought capable of performing the malolactic reaction. We characterized this, as well as the species tolerance for the stressors of the wine environment. Finding it unable to survive in wine, we produced a fully circularized genome of O. alcoholitolerans and performed a comparative genomics analysis to identify the O. oeni genes that enable it to tolerate the pH and ethanol, which O. alcoholitolerans and O. kitaharae lacks. In conclusion, we have used the new technologies of next generation sequencing to produce high-quality genomes and performed extensive, species-wide comparative analyses that allowed us to identify patterns in gene presence that provide likely explanations for environmental adaptation.

京都大学 / 0048 / 新制・課程博士 / 博士(農学) / 甲第22485号 / 農博第2389号 / 新制||農||1075(附属図書館) / 学位論文||R2||N5265(農学部図書室) / 京都大学大学院農学研究科応用生物科学専攻 / (主査)教授 吉田 天士, 教授 澤山 茂樹, 教授 菅原 達也 / 学位規則第4条第1項該当 / Doctor of Agricultural Science / Kyoto University / DFAM

Bottled drinking water is a common form of water consumption that has grown in popularity and dependency. With countless types and brands available, there are factors to consider regarding the variations in mineral content and microbiological quality, and environmental consequences associated with importing natural bottled waters. Saudi Arabia is the largest desalinated water producer, and although there are scarce natural water resources, consuming locally produced water can alleviate environmental pressures, so long as local bottled water is of good quality and provides the basic function of drinking water. The objective of this study is to scrutinize the variations in bottled waters available in the Saudi market and compare local and imported waters regarding water quality and compliance with health regulations. Surveying local bottled waters revealed that over 60% were imported, adding to the environmental significance. Results from inductively coupled plasma-atomic emission spectroscopy, ion chromatography, and pH meters indicate most bottled waters were compliant with health standards. Flow Cytometry (FCM) was used to determine microbial cell densities. Purified waters, on average, contained significantly lower concentrations than other water types, while mineral waters contained the most cells per milliliter. Microbiological stability was examined through incubating samples at 30°C to promote microbial growth, and results were analyzed using an online FCM system. Purified waters had the least amount of growth and to a lesser degree than mineral and sparkling, which varied in microbial growth responses. DNA extractions and microbial community analyses were performed on select mineral waters before and after incubation. Results showed microbial genera found were similar between samples from the same country and some were unique. All samples exhibited some change in microbial community composition after incubation, signifying that certain communities thrived more than others. Proteobacteria was the most common abundant phylum among samples. Although analysis was restricted to the genera level, there were no health concern associated with types of bacteria discovered. In conclusion, local purified waters had greater microbiological quality and stability than imported waters, and mineral compositions were within the healthy range. Reducing reliance on foreign natural waters may help alleviate environmental consequences and improve local economy