Lai Chi-wai. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2001. / Includes bibliographical references (leaves 93-100). / Abstracts in English and Chinese. / ACKNOWLEDGEMENTS --- p.I / ABSTRACT --- p.II / 摘要 --- p.IV / ABBREVIATION --- p.VI / TABLE OF CONTENTS --- p.VII / LIST OF FIGURES --- p.XII / LIST OF TABLES --- p.XIV / Chapter CHAPTER 1 --- INTRODUCTION --- p.1 / Chapter 1.1 --- Catalases --- p.2 / Chapter 1.2 --- Classification of catalases --- p.3 / Chapter 1.2.1 --- Catalase peroxidase (HPI) --- p.3 / Chapter 1.2.2 --- Monofunctional catalases (HPII) --- p.6 / Chapter 1.2.2.1 --- NADPH in catalases --- p.9 / Chapter 1.2.3 --- Mn-catalases --- p.11 / Chapter 1.3 --- Sources and cytotoxic effects of hydrogen peroxide --- p.13 / Chapter 1.4 --- The Chironomidae --- p.14 / Chapter 1.4.1 --- Life cycle of Chironomidae --- p.14 / Chapter 1.4.2 --- Bloodworms --- p.18 / Chapter 1.4.3 --- Sources of bloodworms --- p.19 / Chapter 1.5 --- Aim of the project --- p.22 / Chapter 1.6 --- Application of the project --- p.22 / Chapter CHAPTER 2 --- MATERIALS AND METHODS --- p.24 / Chapter 2.1 --- Protein determination --- p.25 / Chapter 2.2 --- In vitro activity assays --- p.27 / Chapter 2.2.1 --- Catalase activity assay --- p.27 / Chapter 2.2.2 --- Peroxidase activity assay --- p.27 / Chapter 2.3 --- Screening of catalase in different subcellular fractions --- p.28 / Chapter 2.3.1 --- Preparation of mitochondrial fractions --- p.28 / Chapter 2.3.2 --- Preparation of microsomal fractions --- p.29 / Chapter 2.3.3 --- Preparation of cytosolic fractions --- p.29 / Chapter 2.3.4 --- Preparation of post-mitochondrial fractions --- p.29 / Chapter 2.4 --- Purification of post-mitochondrial catalase --- p.29 / Chapter 2.4.1 --- Preparation of post-mitochondrial fractions --- p.30 / Chapter 2.4.2 --- Ethanol-chloroform precipitation --- p.30 / Chapter 2.4.3 --- Affinity chromatography --- p.30 / Chapter 2.4.4 --- Cation exchange chromatography --- p.31 / Chapter 2.5 --- Molecular mass determination --- p.34 / Chapter 2.6 --- Isoelectric focusing --- p.39 / Chapter 2.7 --- Kinetic studies of the purified enzyme --- p.42 / Chapter 2.7.1 --- Optimal pH --- p.42 / Chapter 2.7.2 --- Thermal stability --- p.42 / Chapter 2.7.3 --- Km and Vmax --- p.42 / Chapter 2.7.4 --- Inhibition studies --- p.43 / Chapter 2.7.4.1 --- "3-amino-1,2,4-triazole" --- p.43 / Chapter 2.7.4.2 --- Potassium cyanide and sodium azide --- p.43 / Chapter 2.8 --- Spectroscopic analysis --- p.44 / Chapter 2.8.1 --- Native enzyme --- p.44 / Chapter 2.8.2 --- Denatured enzyme --- p.44 / Chapter 2.8.3 --- Determination of pyridine hemochrome --- p.44 / Chapter 2.9 --- N-terminal amino acid sequence analysis for blotted protein --- p.45 / Chapter 2.9.1 --- Semi-dry electroblotting --- p.45 / Chapter 2.9.2 --- Protein staining on PVDF membrane --- p.46 / Chapter 2.9.3 --- N-terminal amino acid sequence analysis --- p.46 / Chapter 2.9.4 --- N-terminal deblocking of protein bound on PVDF membrane… --- p.47 / Chapter 2.9.5 --- BLAST® search --- p.48 / Chapter CHAPTER 3 --- RESULTS --- p.49 / Chapter 3.1 --- Catalase in different sub-cellular fractions --- p.50 / Chapter 3.2 --- Purification of post-mitochondrial catalase --- p.51 / Chapter 3.2.1 --- Ethanol-chloroform precipitation --- p.51 / Chapter 3.2.2 --- Affinity chromatography --- p.51 / Chapter 3.2.3 --- Cation exchange chromatography --- p.52 / Chapter 3.3 --- Determination of molecular mass --- p.57 / Chapter 3.4 --- Determination of isoelectric point --- p.57 / Chapter 3.5 --- Kinetic studies of the catalase --- p.62 / Chapter 3.5.1 --- Optimal pH --- p.62 / Chapter 3.5.2 --- Thermal stability --- p.62 / Chapter 3.5.3 --- Km and Vmax --- p.65 / Chapter 3.5.4 --- Inhibition studies --- p.65 / Chapter 3.5.4.1 --- "3-amino-1,2,4-triazole" --- p.65 / Chapter 3.5.4.2 --- Potassium cyanide and sodium azide --- p.65 / Chapter 3.5.5 --- Catalase peroxidase activity --- p.66 / Chapter 3.6 --- Spectroscopic analysis --- p.73 / Chapter 3.6.1 --- Native enzyme --- p.73 / Chapter 3.6.2 --- Denatured enzyme --- p.73 / Chapter 3.6.2.1 --- Potassium cyanide --- p.73 / Chapter 3.6.2.2 --- Sodium azide --- p.73 / Chapter 3.6.3 --- Pyridine hemochrome characterization --- p.73 / Chapter 3.7 --- N-terminal amino acid sequence analysis --- p.79 / Chapter CHAPTER 4 --- DISCUSSION --- p.81 / Chapter 4.1 --- Subcellular locations of catalase in bloodworms --- p.82 / Chapter 4.2 --- Purification of post-mitochondrial catalase --- p.82 / Chapter 4.3 --- Physical properties of the purified enzyme --- p.84 / Chapter 4.3.1 --- Native and subunit molecular mass --- p.84 / Chapter 4.3.2 --- Isoelectric point --- p.85 / Chapter 4.4 --- Kinetic properties of the purified enzyme --- p.85 / Chapter 4.4.1 --- Optimal pH --- p.85 / Chapter 4.4.2 --- Thermal stability --- p.85 / Chapter 4.4.3 --- Km and Vmax --- p.87 / Chapter 4.4.4 --- Inhibition studies --- p.87 / Chapter 4.4.5 --- Catalase peroxidase activity --- p.87 / Chapter 4.5 --- Spectroscopic analysis --- p.88 / Chapter 4.5.1 --- Native and denatured enzyme --- p.88 / Chapter 4.5.2 --- Pyridine hemochrome characterization --- p.88 / Chapter 4.6 --- N-terminal amino acid analysis --- p.89 / Chapter 4.7 --- Conclusions --- p.89 / REFERENCES --- p.93

L’évolution du climat Holocène a fait jusqu’ici l’objet de très peu d’études au niveau de la Bretagne. Celles-ci, s’inscrivant dans un contexte à la fois paléo-environnemental et archéologique, présentent souvent un caractère local et ponctuel qui permet difficilement de replacer les changements paléoenvironnementaux observés dans un contexte climatique plus global. Pourtant, la Bretagne, par sa particularité géographique et son exposition océanique, constitue un milieu de prédilection pour l’étude du climat puisqu’elle est fortement exposée, à l’actuel comme ce fut aussi le cas par le passé, aux aléas climatiques typiques de l’Atlantique Nord (régimes de tempêtes notamment) induits par les changements de configurations atmosphériques et océaniques. De plus, la région Bretagne présente une grande diversité de milieux littoraux, avec la présence de rias, d’estuaires, de marais salants, etc. Or, les environnements côtiers sous influence fluviale sont des puits sédimentaires soumis à de forts taux de sédimentation qui offrent l’opportunité d’enregistrer les changements environnementaux passés sur des milliers d’années. Les milieux d’arrière-plage (dunes, lagunes, tourbières), abondamment présents dans la région, constituent ainsi des domaines protégés de l’érosion marine et renferment des remplissages sédimentaires ayant potentiellement enregistré l’évolution séculaire et/ou millénaire du littoral breton, avec très certainement l’empreinte de certains évènements hydrodynamiques majeurs, mais aussi des traces de l’occupation humaine, dont certains auteurs ont montré le caractère précoce et croissant depuis le mésolithique. Ici, nous nous proposons de conduire des reconstitutions paléoenvironnementales et paléoclimatiques à partir de plusieurs séquences sédimentaires prélevées autour du littoral breton, afin de mieux comprendre la réalité des forçages naturels et/ou anthropiques sur les écosystèmes analysés. Pour mener à bien notre étude, nous avons développé une approche paléoécologique multidisciplinaire impliquant plusieurs disciplines complémentaires : sédimentologie et pétrographie, palynologie (pollen et dinokystes) et micropaléontologie(foraminifères benthiques et Chironomidae). Au final, l’analyse des séquences sédimentaires fournit une vision très précise et complète des fluctuations climatiques et environnementales qui ont caractérisé le Nord-Ouest de la France durant la seconde partie de l’Holocène, i.e. depuis les 7000 dernières années. Le recoupement des données entre les différents sites étudiés (1 site Nord Finistère et 2 sites Morbihan),montre l’hétérogénéité spatiale des données paléoécologiques qui caractérisent le secteur. Alors que les séquences prélevées sur les côtes sud bretonnes enregistrent un signal d’anthropisation dès le Néolithique moyen, celle-ci n’apparaît qu’à partir de l’Age du Bronze sur les côtes nord bretonnes, en accord avec les données archéologiques. De plus, nos données montrent que la néolithisation ne s’effectue pas de façon continue mais montre des phases de régression autour de la transition Néolithique final - Age du Bronze, en lien très certainement avec des détériorations climatiques. En effet, les périodes de froid signalées sur le nord de l’Europe durant l’Holocène et caractérisées par des récurrences de tempêtes dans la région Bretagne, semblent synchrones avec les chutes des marqueurs d’anthropisation mais aussi avec les retraits des populations de la côte vers l’intérieur des terres tels que décrits par les études archéologiques depuis le Néolithique final. Ces résultats suggèrent un impact direct du climat sur les dynamiques de population et d’occupation des territoires depuis le Néolithique en Bretagne. / No

This thesis revealed Australian lakes to be richer in chironomid (Diptera: Insecta) species than was previously recognised. A methodology for the collection of chironomid exuviae from lakes was developed using a 12-month study of exuviae from a single lake, Lake McKenzie, Jervis Bay. The method is a rapid and effective way to produce an inventory of species living within a lake. In addition, a distinct biogeographical pattern was detected for chironomid species from a survey of chironomids from southern and eastern Australian freshwater lakes. Geographical location of the lake was more influential on the distribution of chironomid species than was the type of lake. The majority of lake dwelling chironomid species in this investigation were restricted to lakes within one of four geographic lake regions; Tasmania, south-eastern Australian mainland, Fraser Island or tropical north Queensland. A temporal investigation of chironomid exuviae was conducted on a pair of small upland Blue Mountain waterways. Abundance and species richness of exuviae exhibited diurnal patterns. A chemical and macro-invertebrate survey of zinc and sewage organic waste discharges to upland streams in the Blue Mountains detected marked ecological impairment. Macro-invertebrate families responded in different ways to the two different types of waste discharge. According to the family-level results, chironomid larvae responded negatively (reduced abundance) to the zinc pollution and positively (increased abundance) to the sewage pollution. Another major finding from this thesis was that chironomid species assemblages, in the streams surveyed, were strongly impaired by zinc-contaminated mine drainage and sewage effluent. This differed to the family-level larval results. This thesis provided the first Australian evidence that many chironomid species are intolerant of heavy-metal pollution. This research also revealed further evidence that many chironomids species are intolerant of sewage pollution. / Doctor of Philosophy (PhD)

This study documents dynamic changes in the mean abundance and body size of arthropods in an ephemeral pond over the course of a summer in order to understand the nature of interactions that shape this community, and to assess the risks posed by exposure to cercariae of the entomopathogenic digenean, Plagiorchis elegans, a potential control agent for larval mosquitoes. Chironomids were the most abundant, comprising almost two-thirds of all arthropods collected; they were followed by chaoborids, the most abundant predator. Other arthropods, in decreasing order of abundance, were the ephemerids, isopods, amphipods, odonates, dytiscids, and hemipterans. Infections were found only in chironomids and were extremely rare and of low intensity, conceivably due to predation of chaoborids on cercariae and may have protected the community from exposure to these parasites. Findings are discussed in terms of using entomopathogenic digeneans as agents in the biological control of mosquitoes.

Made available in DSpace on 2016-06-02T19:31:36Z (GMT). No. of bitstreams: 1 DissCSNO.pdf: 5347163 bytes, checksum: bb3325e5f0f024bc489c3711209e89bb (MD5) Previous issue date: 2006-02-24 / The present work aimed to study the morphology, taxonomy and bionomy of Ablabesmya Johannsen, 1905 (Diptera, Chironomidae, Tanypodinae) species of the State of São Paulo. This genus of world distribution prise 69 species, of which ten are registered in the Neotropical region, however none specifically mentioned for Brazil. The analyzed material derives from three lotic systems (streams and rivers) and seven lentic systems (reservoirs and lakes), which are located in three cities of the State of São Paulo (São Carlos, Luís Antônio and Brotas). The sampling followed the conventional methods of benthonic studies. The larvae were kept alive in the laboratory to obtain the associations between larva, pupa and adult. The identification of the species was based on the original descriptions. The results allowed the identification of eight species, seven of wich are new species and the second record of Ablabesmyia metica Roback (1983) for the Neotropical region and the first one to Brazil, describes the immature stages, unknown before. The new species are described with the respective diagnoses. The ecologic information on the genus are discussed and perspectives for the future studies are mencioned. / O objetivo deste trabalho foi estudar a morfologia, taxonomia e bionomia das espécies de Ablabesmyia Johannsen, 1905 (Diptera, Chironomidae, Tanypodinae) do Estado de São Paulo. Este gênero de distribuição mundial compila 69 espécies, das quais dez são registradas para a região Neotropical, entretando nenhuma citada especificamente para o Brasil. O material analisado tem como procedência três sistemas lóticos (rios e córregos) e sete sistemas lênticos (represas e lagoas), os quais estão situados em três municípios do Estado de São Paulo (São Carlos, Luís Antônio e Brotas). As coletas seguiram a metodologia convencional para estudos bentônicos. As larvas foram mantidas vivas em laboratório até a obtenção das três fases do desenvolvimento; larva, pupa e adulto. As identificações das espécies foram baseadas nas descrições originais. Os resultados permitiram a identificação de oito espécies, das quais sete são espécies novas para o gênero e o segundo registro de A. metica ROBACK (1983) para a região Neotropical e o primeiro do Brasil, onde há a descrição dos estágios imaturos, anteriormente desconhecidos. As novas espécies são descritas com as respectivas diagnoses. As informações ecológicas do gênero são discutidas e perspectivas para trabalhos futuros são mencionadas.

The macroscopic bottom fauna of Marion Lake, British Columbia, was investigated from May, 1963 to September, 1964. The primary object was to document seasonal differences within and between specific populations and to interpret these difference in terms of the life histories of the species and their availability to a predator, rainbow trout. A detailed examination of the 51 species of Chironomidae known to occur in the lake has shown that the ecological characteristics of many of the closely related species are very different. Small species generally fed primarily on phytoplankton while the larger species ate more detritus and organic debris or other invertebrates. Predaceous species had more uniform distributions in the lake than did herbivores or detritus feeders. Most species of the subfamily Orthocladinae emerged in spring or fall and grew rapidly during the winter. The species of Chironominae and Tanypodinae usually emerged during the spring and summer and grew very little during the winter. Summer emerging Chironominae emerged later over deep water whereas the Tanypodinae emerged at much the same time above all depths. Larvae which underwent frequent vertical migrations had a higher mortality rate and were found more frequently in the stomachs of rainbow trout than larvae which rarely or never migrated. Many of the benthic organisms were not effectively utilized by rainbow trout. Organisms which numerically constituted approximately 1.5% of the benthic fauna accounted for more than 50% of the food items found in the trout stomachs. Large forms, such as the Odonata and Trichoptera and species which frequently moved off the bottom, formed the bulk of the food; small species and some of the species which were entirely benthic were rare or absent in the trout stomachs analyzed. This study has shown some of the advantages of working at the species level. Although studies at this level are tedious and often necessitate a time consuming taxonomic study, the additional information is likely to justify the extra effort. The results of ecological investigations that are not carried out at the species level ignore, or at best oversimplify, relationships that exist. Indeed, a comprehensive understanding of, for example, energy transfer within a community, is impossible without a detailed knowledge of the life histories of the species involved. / Science, Faculty of / Zoology, Department of / Graduate

No description available.

Un futur climat planétaire plus chaud avec une sècheresse plus marquée durant toute l’année est aujourd’hui prévus, en particulier pour les hautes latitudes de l’hémisphère nord. Ces changements environnementaux seraient marqués par des fréquences et des intensités plus importantes des perturbations naturelles, qui pourraient menacer l’intégrité de certains écosystèmes forestiers boréaux. L’une des conséquences prévisibles serait une augmentation significative de l’occurrence des feux de forêt qui est déjà et deviendrait davantage encore l’élément perturbateur majeur de ces écosystèmes. Le devenir des écosystèmes boréaux suscite donc de nombreuses interrogations et des incertitudes, ce qui amène à souligner la nécessité d’une bonne caractérisation des changements climatiques spatio-temporels et de leurs conséquences sur ces écosystèmes.Afin de maintenir ces paysages forestiers dans les limites de leur variabilité naturelle, le Ministère des Forêts, de la Faune et des Parcs du Québec (MFFP) se propose, dans un contexte d’aménagement écosystémique durable, d’établir un Registre des états de référence. Dans cette perspective, l’objectif de recherche de cette thèse était de produire des reconstitutions des paléotempératures holocènes les plus robustes possibles au sein de la pessière à mousses de l’ouest du Québec. La recherche a porté sur une approche multi-indicateurs incluant les Chironomidae et le pollen contenus dans les sédiments des deux lacs retenus. Les résultats mettent en évidence la pertinence des capsules céphaliques de Chironomidae pour reconstituer à haute résolution temporelle, les paléotempératures holocènes du mois d’août provenant du Lac Aurélie à l’aide de deux Fonctions de Transfert (FT). La FT Est canadienne s’est avérée plus en accord avec le signal régional que la FT canadienne. De surcroit, les assemblages de Chironomidae du Lac Lili ont donné une reconstitution surestimée des paléotempératures en raison de la faible profondeur de ce lac. Enfin, la reconstitution quantitative combinée (Chironomidae et pollen) des paléotempératures du mois d’août a souligné trois périodes principales: (i) 8500-4500 ans AA, identifiée comme le Maximum Thermique de l’Holocène (HTM) durant laquelle les températures d’août sont plus chaudes que les températures actuelles; (ii) 4500-1000 ans AA, marquée par le début de la période Néoglaciaire froide, où les températures oscillent autour des valeurs actuelles et (iii) les derniers 1000 ans AA, caractérisés par une diminution générale des températures. Plusieurs événements climatiques courts ont également été identifiés pour chaque période : (i) l’évènement froid de 8200 ans AA; (ii) l’évènement chaud de 4200 ans AA; (iii) la période chaude dite Romaine (RWP, 1900-1700 ans AA) ; (iv) la période froide de l’Âge Sombre (DACP, 1700-1500 ans AA) ; (v) l’Anomalie Climatique Médiévale (MCA, 1100 ans AA) et (vi) le Petit Âge Glaciaire (LIA, 500-250 ans AA). Une série d’événements froids est également observée vers 5900, 4300, 2800 et 400 ans AA, se référant probablement aux évènements « Bond ». La durée et l’amplitude observées des événements climatiques rejoignent les enregistrements régionaux, nord-américains et plus largement ceux de l’Hémisphère Nord. Par ailleurs, la synthèse soulignant les relations entre climat-végétation-feu confirme l’influence d’une instabilité climatique sur les changements des régimes des feux, principalement causée par les variations saisonnières des précipitations et de l'irrégularité des épisodes de sécheresse. Toutefois, notre reconstitution combinée des paléotempératures indique que les grands feux coïncident avec l’évènement ponctuel de 4200 ans AA et ceux survenus à partir de 2000 ans (RWP, DACP, MCA et LIA). La température estivale conjuguée à d’autres variables climatiques (sécheresse estivale, ensoleillement estival, et précipitations annuelles) joue donc un rôle important dans la variation du régime des feux à l’ouest du Québec. / A future warmer global climate with more severe annual drought has been predicted, especially for the high latitudes of the Northern Hemisphere. Such an environmental change would be marked by higher frequencies and intensities of natural disturbances, which could threaten the integrity of some boreal forest ecosystems. One of the foreseeable consequences would be a significant increase in the occurrence of wildfires, which is already the major disruptive element of these ecosystems. The future of boreal ecosystems thus raises many questions and uncertainties, highlighting the need for a good characterization of spatio-temporal climate changes and its consequences on these ecosystems.In order to maintain these forest landscapes within the limits of their natural variability, the Ministère des Forêts, de la Faune et des Parcs du Québec (MFFP) proposes, in a context of sustainable ecosystem management, to establish a registry of reference states. In this perspective, the research objective of this thesis was to produce the most possible robust reconstructions of Holocene temperatures within the spruce-moss forest of western Québec. The research focused on a multi-indicator approach including Chironomidae and pollen deposited in the sediments of two selected lakes.The results highlight the relevance of Chironomidae head capsules to reconstruct at high temporal resolution the August Holocene temperatures from Lac Aurélie using two Transfer Functions (TF). The East Canadian TF was more in agreement with the regional signal than the Canadian TF. In addition, chironomid assemblages from Lac Lili gave an overestimated paleotemperature reconstruction due to the specific shallowness of this lake. Finally, the combined quantitative reconstruction (Chironomidae and pollen) of the August palaeotemperatures suggested three distinct periods: (i) 8500-4500 years BP, identified as the Holocene Thermal Maximum (HTM) during which the August temperatures are warmer than modern temperatures; (ii) 4500-1000 cal BP, marked by the beginning of the cold Neoglacial period, where the temperatures oscillate around the current values and (iii) the last 1000 cal BP, characterized by a general decrease of the temperatures. Several short climatic events were also identified for each period: (i) the 8200 cal BP cold event; (ii) the 4200 cal BP warm event; (iii) the Roman Warm Period (RWP, 1900-1700 cal BP); (iv) the Dark Age Cold Period (DACP, 1700-1500 cal BP); (v) Medieval Climate Anomaly (MCA, 1100 cal BP) and (vi) the Little Ice Age (LIA, 500-250 cal BP). A series of cold events is also observed around 5900, 4300, 2800 and 400 cal BP, probably corresponding to the "Bond" events. The observed duration and amplitude of climate events are consistent with regional, North American records, and more widely those of the Northern Hemisphere. In addition, the synthesis highlighting climate-vegetation-fire relationships confirms the influence of climatic instability on changes in fire regimes mainly driven by seasonal variations in rainfall and frequency of drought events. However, our paleotemperatures combined reconstitution indicated that large fires coincide with the occasional event of 4200 cal BP and those occurring from 2000 cal BP onwards (RWP, DACP, MCA and LIA). Summer temperatures combined with other climatic variables (summer drought, summer insolation, and annual precipitation) play an important role in the variation of the fire regime in western Québec.

As comunidades bentônicas são relevantes nos ecossitemas aquáticos devido ao seu papel no funcionamento dos ecossistemas e também como mercadores ou indicadores de paleocondições. Sua distinção, similarmente à de outras comunidades, estão condicionadas por fatores abióticos e pela interação entre eles, determinando a estrutura da comunidade que se estabelecerá. Pelo presente trabalho objetivou-se determinar a composição taxonômica, a abundância e a distribuição espacial da comunidade bentônica, amostrando-se em 20 pontos na lagoa do Caçó. Um total de 14 pontos foram estabelecidos na região litorânea (7 na margem esquerda e 7 na margem direita) e 6 na região limnética. A variação sazonal em parâmetros da comunidade e também das características físicas e químicas da água e do sedimento foram analisadas e relacionadas aos deslocamentos da Zona de Convergência Intertropical. A lagoa do Caçó está localizada ao Norte do Estado do Maranhão e possui aproximadamente 5 km de comprimento e largura média de 500 metros. As coletas foram realizadas em abril (período chuvoso) e novembro (período seco) de 1999. Pelos resultados obtidos, ficam evidentes as diferenças tanto na riqueza de táxons quanto na diversidade de espécies e na densidade das populações em função da sazonalidade. A diversidade, a riqueza e a densidade foram maiores no período seco do que no período chuvoso. A família Chironimidae (Diptera Insecta) foi o mais importante componente da comunidade, representando aproximadamente 50% do total da fauna bentônica da lagoa. O sedimento da Lagoa do Caçó tem baixo conteúdo de matéria orgânica, sendo classificado como mineral. A análise da composição granulométrica mostrou que o sedimento é heterogêneo, sendo arenoso na margem esquerda, predominantemente argiloso na margem direita e limo arenoso e argiloso na região limnética. Em relação às variáveis limnológicas, observaram-se microestratificações térmicas e ) químicas. Os índices tróficos aplicados permitiram classificar a lagoa do Caçó como oligotrófico. A alternância dos períodos seco e chuvoso atuou como a principal função de força condicionado as alterações nas condições físicas e químicas da água bem como na comunidade bentônica no ano de 1999. Além disso, a identificação dos Chironomidae atuais da lagoa do Caçó até o nível de gênero, e o fato deste grupo produzir remanescentes fossilizáveis os tornam bons marcadores paleolimnológicos e paleoclimáticos que podem ser utilizados para inferir sobre as condições lacustres e climáticas passadas. / Benthic communities are relevant in aquatic ecosystems due to their role in the ecosystem functioninq and also as rnarkers or indicators of paleoconditions. Their distribution, similarly to other communities, is conditioned by abiotic and biotic factors and to their interactions, which determine the structure of the community that will be established. The present work aimed to determine the taxonomic composition, the abundance and spatial distribution of the benthic community, by collecting at 20 sampling points in Caçá Lake. A total of 14 points were located in the littoral region (7 on the right margin and 7 on the left one) and 6 in the limnetic zone. Seasonal variation on community parameters and also of chemical and physical characteristics of water and sediment were analysed and related to the seasonal displacement of the Intertropical Convergence Zone. Caçó Lake is located at the North of Maranhão State and it has approximately 5 km of length and 500m mean width. Sampling was carried out in April (rainy season) and November (dry season). The results evidenced that there are differences in the richness of taxa, diversity and density of benthic species that are mainly related to the seasonality. The diversity, richness and density were higher in the dry than in the rainy period. The family Chironomidae (Diptera, Insecta) was the most important component of the community representing approximately 50% of the total benthic fauna of the lake. The sediment of Caçá Lake has low organic content being classified as a mineral one. The granulometric composition shows that the sediment is heterogeneous being sandy at the left margin, clayey in the right margin and sandy-clayey in the limnetic zone. In relation to the limnological variables, thermal and chemical (pH, dissolved oxygen and temperature) micro-stratifications were observed for both periods sampled. The trophic state indices applied have allowed to classifv Lake Caçá as an oligotrophic lake. The alternation of dry and rainy periods can be considered the main forcing function causing the changes in physical and chemical water characteristics as well on benthic community in the year 1999. The identification of the present Chironomidae at the genus level and the fact that this group does produce fossilizing remains make them good paleolimnological and paleoclimatic tracers that may be used to infer on past lake and climate conditions.

Chong, King Wai Isaac. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2010. / Includes bibliographical references (leaves 99-104). / Abstracts in English and Chinese. / Acknowledgements --- p.i / Abstract --- p.ii / 論文摘要 --- p.iv / Table of Contents --- p.vi / Lists of Figures --- p.ix / List of Tables --- p.xi / List of Abbreviations --- p.xii / Chapter Chapter One: --- Introduction --- p.1 / Chapter 1.1 --- Overview of Glyceraldehyde 3-phosphate Dehydrogenases --- p.1 / Chapter 1.2 --- Properties And Molecular Structures of GAPDH --- p.3 / Chapter 1.3 --- Action Mechanism of GAPDH --- p.6 / Chapter 1.4 --- Novel Functions of GAPDH Unrelated to Carbohydrate Metabolism --- p.8 / Chapter 1.5 --- Effects of Heavy Metal on Enzyme Activity And Gene Expression of GAPDH --- p.10 / Chapter 1.6 --- Metal Binding Properties And Metal Binding Sites of GAPDH --- p.12 / Chapter 1.7 --- Isolation And Purification of GAPDH from Different Organisms --- p.13 / Chapter 1.8 --- Development of New Purification Method of GAPDH Using Immobilized Metal Affinity Chromatography --- p.15 / Chapter 1.9 --- Study of GAPDH from Chironomidae Larvae --- p.16 / Chapter 1.10 --- Aims of Study --- p.18 / Chapter Chapter Two: --- Methods And Materials --- p.19 / Chapter 2.1 --- Isolation of Native Chironomidae GAPDH --- p.19 / Chapter 2.1.1 --- Chemicals And Reagents --- p.19 / Chapter 2.1.2 --- Reagents --- p.19 / Chapter 2.1.3 --- Preparation of Crude Protein Extract from Chironomidae Larvae --- p.24 / Chapter 2.1.4 --- Immobilized Metal Affinity Chromatography --- p.24 / Chapter 2.1.5 --- Large Scale Preparation of Crude Protein Extract --- p.25 / Chapter 2.1.6 --- Ammonium Sulfate Fractionation --- p.25 / Chapter 2.1.7 --- Copper Affinity Column Chromatography --- p.26 / Chapter 2.1.8 --- Dye Affinity Column Chromatography --- p.26 / Chapter 2.2 --- Identification of Chironomidae GAPDH --- p.27 / Chapter 2.2.1 --- Chemicals And Reagents --- p.27 / Chapter 2.2.2 --- Reagents --- p.28 / Chapter 2.2.3 --- Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis --- p.32 / Chapter 2.2.4 --- Non-Denaturing Polyacrylamide Gel Electrophoresis --- p.33 / Chapter 2.2.5 --- Protein Extraction from Coosmassie Blue Stained Polyacrylamide Gel --- p.33 / Chapter 2.2.6 --- N-terminal Amino Acid Analysis --- p.33 / Chapter 2.2.7 --- Sequence Analysis --- p.34 / Chapter 2.3 --- Kinetic Analysis of Chironomidae GAPDH --- p.34 / Chapter 2.3.1 --- Chemcials And Reagents --- p.34 / Chapter 2.3.2 --- Reagents --- p.34 / Chapter 2.3.3 --- Determination of Enzyme Concentration And GAPDH Activity --- p.35 / Chapter 2.4 --- Molecular Cloning of Chironomidae GAPDH --- p.36 / Chapter 2.4.1 --- Chemicals And Reagents --- p.36 / Chapter 2.4.2 --- Reagents --- p.37 / Chapter 2.4.3 --- RNA Extraction from Chironomidae Larvae --- p.41 / Chapter 2.4.4 --- DNase I Removal of Genomic DNA Contamination --- p.42 / Chapter 2.4.5 --- RNA Concentration Determination And RNA Agarose Electrophoresis --- p.42 / Chapter 2.4.6 --- First Strand cDNA Synthesis --- p.43 / Chapter 2.4.7 --- pRSet A B C Vectors --- p.43 / Chapter 2.4.8 --- Cloning Primer Design --- p.45 / Chapter 2.4.9 --- Polymerase Chain Reaction --- p.45 / Chapter 2.4.10 --- DNA Agarose Electrophoresis --- p.46 / Chapter 2.4.11 --- Restriction Enzyme Digestion of Insert And Plasmid --- p.46 / Chapter 2.4.12 --- Ligation of Plasmid And Insert DNA --- p.46 / Chapter 2.4.13 --- Preparation of Chemically Competent E. coli --- p.47 / Chapter 2.4.14 --- Transformation of Plasmid by Heat Shock --- p.47 / Chapter 2.4.15 --- Colony PCR --- p.48 / Chapter 2.5 --- Recombinant Protein Expression And Purification --- p.48 / Chapter 2.5.1 --- Chemicals And Reagents --- p.48 / Chapter 2.5.2 --- Reagents --- p.49 / Chapter 2.5.3 --- Protein expression by IPTG --- p.51 / Chapter 2.5.4 --- Protein purification by Nickel Affinity Column Chromatography --- p.52 / Chapter 2.5.5 --- EnterokinaseMax ´ёØ Removal of Polyhistidine Fusion Tag --- p.52 / Chapter 2.5.6 --- Western Blotting of Protein --- p.53 / Chapter Chapter Three: --- Results --- p.54 / Chapter 3.1 --- Two Affinity Chromatography Methods for GAPDH Purification --- p.54 / Chapter 3.2 --- Isolation And Purification of Native Chironomidae GAPDH --- p.54 / Chapter 3.3 --- Identification of Chironomidae GAPDH --- p.62 / Chapter 3.3.1 --- N-terminal amino acid analysis --- p.62 / Chapter 3.3.2 --- Sequence Analysis --- p.62 / Chapter 3.4 --- Molecular Cloning of Chironomidae GAPDH --- p.63 / Chapter 3.5 --- Isolation And Purification of recombinant Chironomidae GAPDH --- p.70 / Chapter 3.6 --- Protein Gel Electrophoresis Analysis of GAPDHs --- p.74 / Chapter 3.7 --- "Effects of Heavy Metals, pH And Temperature on GAPDHs" --- p.76 / Chapter 3.7.1 --- Heavy Metal Effect --- p.76 / Chapter 3.7.2 --- pH Effect --- p.76 / Chapter 3.7.3 --- Temperature --- p.77 / Chapter 3.8 --- Kinetic Analysis of GAPDHs --- p.84 / Chapter Chapter Four: --- Discussion --- p.89 / Chapter 4.1 --- New Method for The Isolation and Purification of Chironomidae GAPDH --- p.89 / Chapter 4.2 --- "Effects of Heavy Metals, pH And Temperature on GAPDHs" --- p.91 / Chapter 4.3 --- Kinetic Analysis of GAPDHs --- p.91 / Chapter 4.4 --- Zinc Activation of Chironomidae GAPDH --- p.92 / Chapter 4.5 --- Future Study --- p.93 / Chapter 4.5.1 --- Sequence Analysis Using Prediction Programmes --- p.94 / Chapter 4.5.2 --- Protein Crystallization --- p.95 / Chapter 4.5.3 --- Site-Directed Mutagenesis --- p.95 / Chapter 4.5.4 --- Biacore Surface Plasmon Resonance --- p.95 / Chapter Chapter Five: --- Conclusion --- p.98 / Chapter Chapter Six: --- References --- p.99