Mesidotea entomon inhabiting Pauline Cove, YT, is larger in total body length at sexual maturity than M. entomon inhabiting Dolomite Lake, NWT. The sex ratio, diet, habitat distributions, size frequency distributions, and fecundity in the two populations are compared. Total carbohydrate, protein, lipid, chitin, and ash contents of both isopods' life stages are compared. Isopods in the cove show reduced carbohydrate and elevated lipid levels. In the 1975, 1978, and 1979 summers, isopods in the cove were internally infected with an extracellular protozoan parasite; those in the lake were uninfected. The infection is systemic. The prevalence of the protozoan infection increases with increasing host size. The host response involved phagocytosis and encapsulation. Transmission may be transovarially and/or by ingestion of the infective stage. The protozoan also infected Mesidotea sibirica. The infection is related to growth, abundance, habitat, reproduction, and biochemical variables.

Deep-water prawns Haliporoides triarthrus and langoustines Metanephrops mozambicus are endemic to the South West Indian Ocean (SWIO) region and make up the largest proportion of deep-water crustacean trawl catches in Mozambique and South Africa. Despite their economic importance to these fisheries, little is known about their distribution, biology and genetic population structure. The metapopulation genetic variation of H. triarthrus and M. mozambicus was assessed from 220 specimens per species collected from three sites in Mozambique (Bazaruto A, Boa Paz and Inhaca), two sites in western Madagascar (Morombe and Tulear) and one site in eastern South Africa (Durban). Two fragments of the mitochondrial region were amplified using universal primers ribosomal 16S subunit (16S) and mitochondrial cytochrome oxidase subunit I (COI). From H. triarthrus, fragments of 569 base pair (bp) (16S) and 1300 bp (COI) were amplified. A total of 207 sequences (16S) and 151 sequences (COI) were recovered, and 69 and 78 haplotypes identified, respectively. Metanephrops mozambicus 16S and COI genes produced similar fragment lengths, and 112 (16S) and 127 haplotypes (COI) were recovered. Both species demonstrated high genetic diversity and significant population differentiation in the SWIO region. Two sister-species (or subspecies) of H. triarthrus were identified, one occurring along the African continental shelf and the other off western Madagascar. Furthermore, individual populations making up each lineage were genetically structured, as indicated by the absence of shared haplotypes, and should be recognized as demographically distinct subspecies. Both species have undergone recent population expansions, likely since the late Pleistocene. The large anti-cyclonic and cyclonic eddies prevalent in the Mozambique Channel, and the boundary area between these eddies and upper Agulhas Current are likely factors driving larval retention or return process, thus giving rise to the observed genetically structured populations. The findings from this study are unique for the SWIO region, and may lead to a paradigm shift in the way that deep-water crustacean stocks are perceived by fisheries managers – instead of single shared stocks, they comprise of many isolated ones, in spite of the dispersal potential of larvae in strong ocean current regimes. Thus stocks should be managed as small independent units. / Thesis (M.Sc.)-University of KwaZulu-Natal, Durban, 2013.

Submitted by Irene Nascimento (irene.kessia@ufpe.br) on 2017-04-18T18:14:43Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) PAIVA, R.J.C..pdf: 5468652 bytes, checksum: 3d8cfedf8656ac46cc27d4c55547d6fc (MD5) / Made available in DSpace on 2017-04-18T18:14:43Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) PAIVA, R.J.C..pdf: 5468652 bytes, checksum: 3d8cfedf8656ac46cc27d4c55547d6fc (MD5) Previous issue date: 2016-07-27 / FACEPE / Atualmente são conhecidas 492 espécies da Família Cirolanidae. Foi realizada uma revisão taxonômica das espécies que ocorrem no oeste do oceano Atlântico (totalizando 66), e uma análise distribucional utilizando o Método de Análise Parcimônia de Endemismo (PAE). O material se encontra depositado nas coleções de carcinologia do Museu de Oceanografia da Universidade Federal de Pernambuco, Museu Nacional do Rio de Janeiro e no Museu de Zoologia da USP. Com o estudo dessas amostras foi possível propor nova chave de identificação, atualizando as já existentes para o Oceano Atlântico, incluindo as espécies descritas mais recentemente. Os exemplares estudados foram dissecados e desenhados com auxílio de microscópio e estereomicroscópio com câmara clara acoplada. Os exemplares foram corados em negro de clorazol e suas peças montadas em lâminas semi-permanentes com gelatina glicerinada. Os desenhos digitais foram confeccionados no programa gráfico Corel Draw®. Foram encontradas oito espécies distribuídas em seis gêneros, sendo dois registros novos, um do gênero Bathynomus (B. miyarei) e outro do gênero Calyptolana (C. hancocki). Foram resistradas seis espécies novas para ciência: duas de Cirolana sp. nov. A, C. sp. nov.B, Dolicholana brucei, D. sp. nov., Natatolana sp. nov., Politolana sp. nov. Dentre as espécies estudadas B. miyarei, C. sp. nov. A, C. sp. nov. B, Dolicholana brucei, D. sp. nov., Natatolana sp. nov., Politolana sp. nov. até o momento são endêmicas para o Brasil. Com base nos dados levantados nessa pesquisa foram descritas seis espécies novas para a costa Brasileira, incluindo regiões de água rasa e mar profundo. Além disso, foram registrados pela primeira vez o gênero Dolicholana Bruce, 1986. A análise de parcimônia de endemismo revelou quatro áreas de endemismo no Atlântico Ocidental, corroborando em parte com algumas propostas já bem estabelecidas na literatura para outros grupos de crustáceos. / There are known 492 species of Family Cirolanidae. It was performed a taxonomic revision of the species occurring in the western Atlantic Ocean (totaling 66), and a distributional analysis using the Parsimony Analysis of Endemicity (PAE). The material analyzed is deposited at carcinology collections of the Oceanography Museum on the Universidade Federal de Pernambuco, National Museum of Rio de Janeiro and on the USP Zoology Museum. With the study of these samples was possible to propose new identification key updating the one which exist to the Atlantic Ocean, including the described species recently. The specimens studied were dissected and drawn with the aid of a microscope and stereo microscope with camera lucida attached. The specimens were stained with dark clorazol and these parts were mounted on semi-permanent slides with glycerinated gelatin. The digital drawings were made in Corel Draw® graphics program. We found eight species distributed in six genera, two new records, a genus of Bathynomus (B. miyarei) and other genus of Calyptolana (C. hancocki). It was registered six new species to science: Two Cirolana sp. nov. A, C. sp nov. B, Dolicholana brucei, D. sp. nov., Natatolana sp. nov., Politolana. sp. nov. Among the species studied B. miyarei, C. sp.nov. A, C. sp. nov. B, Dolicholana brucei, D. sp. nov., Natatolana sp. nov., Politolana sp. nov. are endemics to Brazil up to now. Based on data collected in this study were described six new species to the Brazilian coast, including areas of shallow water and deep sea. Besides that, it was registered to the first time a genus of Dolicholana Bruce, 1986. The parsimony analysis of endemicity revealed four areas of endemicity in the western Atlantic, partly corroborating with some proposals already well established in the literature for others crustacean groups.

Die Keimstreifverlängerung der Malacostraca wird durch ein festgelegtes Zellteilungsmuster der posterioren Teilungszone erreicht. Dort geben anfangs in einem Halbkreis angeordnete Ektoteloblasten ihre Tochterzellen nach anterior ab, welche das für malakostrake Krebse typische Gittermuster ausbilden. Dabei spielen die Ektoteloblasten eine besondere Rolle, da sie das Zellmaterial für die anschließende Segmentierung produzieren. Im Gegensatz zu Vertretern basal abzweigender Taxa der Malacostraca, bei denen sich der Halbkreis der Ektoteloblasten aus zwei Halbreihen oder aus einem Zellring bildet, sind bei den Isopoden, zu denen auch die Kellerassel Porcellio scaber zählt, die Ektoteloblasten von Beginn an in einem Halbkreis angeordnet. Das Teilungsmuster bei der Verlängerung des Keimstreifens wurde bereits bei vielen Vertretern der Malacostraca, einschließlich P. scaber, ausführlich untersucht. Daten über die Bildung der Ektoteloblasten und den Beginn ihrer Teilungsaktivität fehlen dagegen weitestgehend. Im Rahmen der vorliegenden Arbeit wurde daher mit Hilfe von Zellmarkierungsversuchen mit Vitalfarbstoffen (wie z.B. Alexa Fluor 488 und Tetramethylrhodamin) und der 4D-Mikroskopie die Bildung der Keimscheibe, die Gastrulation und der Beginn der Keimstreifverlängerung bei P. scaber untersucht. Ein besonderer Fokus lag dabei auf den Ektoteloblasten und den umliegenden Zellen, deren Zelllinien untersucht wurden. Wie in der vorliegenden Arbeit gezeigt werden konnte, sind (i) die Ektoteloblasten bereits vor Beginn der Gastrulation in einem äußeren Halbkreis formiert, (ii) zwei Zellen, die Teile der späteren Mittellinie bilden, bis kurz vor Beginn der Teilungstätigkeit der Ektoteloblasten in deren Reihe eingereiht, (iii) die Ektoteloblasten erst mit der Fertigstellung der ersten e-Reihe als große, verlängerte Zellen erkennbar und es ist (iv) ein zweiter Halbkreis vorhanden, der sich ausschließlich in posteriore Richtung teilt und dadurch den Gastrulationsporus schließt. / In malacostracan crustaceans, the elongation of the germ band is realized by an invariant cell division pattern of the posterior growth zone. The ectoteloblasts are arranged in a semicircle and divide in anterior direction in a fixed pattern. Their progeny are arranged in the typical grid pattern of malacostracan crustaceans and provide the cell material for later segmentation processes. In contrast to other malacostracans, where the ectoteloblast appear in two half rows or a cell ring, the ectoteloblasts are arranged in a semicircle in isopods from the outset. The cell division pattern during elongation of the germ band is well studied in many malacostracan species including the isopod Porcellio scaber. However, details about formation of the ectoteloblasts and beginning of their cell division activity are missing. Therefore, the aggregation of the germ disc, the gastrulation, and the beginning of the elongation of the germ band in Porcellio scaber were examined using vital dyes (e.g. Alexa Fluor 488 and Tetramethylrhodamine) in cell labeling experiments combined with 4Dmicroscopy. The formation and cell-lineages of the ectoteloblasts and their surrounding cells were analyzed. The results suggest that (i) the ectoteloblasts are arranged in a semicircle before gastrulation starts, (ii) two cells forming part of the arising midline are lined up with the ectoteloblasts until they start their cell division activity in a predetermined pattern, (iii) the differentiation of ectoteloblasts is discernable after formation of the first e-row when they appear as big and elongated cells and (iv) a second semicircle exists whose cells divide in posterior direction and, consequently, enclose the gastrulation pore.

No description available.

Chan Kwan-ling. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (leaves 91-105). / Abstracts in English and Chinese. / Abstract --- p.i / Acknowledgements --- p.v / Contents --- p.vi / List of figures and tables --- p.ix / Chapter Chapter 1 --- Introduction / Chapter 1.1 --- Literature review --- p.1 / Chapter 1.1.1 --- The effect of pollutants on the genetic materials of aquatic organisms --- p.1 / Chapter 1.1.1.1 --- Response of individual to genotoxicants --- p.1 / Chapter 1.1.1.2 --- Effects of genotoxicants on population structure --- p.3 / Chapter 1.1.2 --- Application of genetic markers in monitoring water pollution --- p.3 / Chapter 1.1.2.1 --- DNA adduct --- p.4 / Chapter 1.1.2.2 --- Sister chromatid exchange (SCE) test --- p.5 / Chapter 1.1.2.3 --- Micronucleus --- p.6 / Chapter 1.1.2.4 --- DNA strand breaks --- p.7 / Chapter 1.1.3 --- Single-cell gel electrophoresis (comet) assay --- p.9 / Chapter 1.1.4 --- Test organisms for comet assay --- p.12 / Chapter 1.2 --- Objective of the present study --- p.13 / Chapter Chapter 2 --- Genotoxicity of pollutants on Hyale crassicornis / Chapter 2.1 --- Introduction --- p.22 / Chapter 2.2 --- Materials and methods --- p.24 / Chapter 2.2.1 --- Sampling of amphipods --- p.24 / Chapter 2.2.2 --- Acclimation --- p.24 / Chapter 2.2.3 --- Acute toxicity test --- p.26 / Chapter 2.2.4 --- The effect of test duration on DNA damage --- p.27 / Chapter 2.2.5 --- Effect of toxicants on DNA damage --- p.28 / Chapter 2.2.6 --- Comet assay --- p.29 / Chapter 2.2.7 --- Chemicals --- p.34 / Chapter 2.2.8 --- Data analysis --- p.34 / Chapter 2.3 --- Results --- p.34 / Chapter 2.4 --- Discussion --- p.47 / Chapter Chapter 3 --- Genotoxicity of hydrogen peroxide on different tissue types of Metapenaeus ensis / Chapter 3.1 --- Introduction --- p.54 / Chapter 3.2 --- Materials and Methods --- p.57 / Chapter 3.2.1 --- Collection and acclimation of shrimps --- p.57 / Chapter 3.2.2 --- Incubation --- p.59 / Chapter 3.2.3 --- Comet Assay --- p.60 / Chapter 3.2.4 --- Chemicals --- p.61 / Chapter 3.2.5 --- Data analysis --- p.61 / Chapter 3.3 --- Results --- p.61 / Chapter 3.4 --- Discussion --- p.67 / Chapter Chapter 4 --- Genotoxicity of wastewater on Hyale crassicornis / Chapter 4.1 --- Introduction --- p.71 / Chapter 4.2 --- Materials and Methods --- p.72 / Chapter 4.2.1 --- Collection of wastewater samples --- p.72 / Chapter 4.2.2 --- Metal content analysis --- p.73 / Chapter 4.2.3 --- Genotoxic effect of wastewater samples on Hyale crassicornis --- p.74 / Chapter 4.2.4 --- Chemicals --- p.76 / Chapter 4.2.5 --- Statistical analysis --- p.77 / Chapter 4.3 --- Results --- p.77 / Chapter 4.3.1 --- Metals content in water samples --- p.77 / Chapter 4.3.2 --- DNA damage --- p.79 / Chapter 4.4 --- Discussion --- p.79 / Chapter Chapter 5 --- Conclusions --- p.89 / References --- p.91

O presente estudo foi realizado ao longo de uma linha de costa, incluindo duas enseadas em Ubatuba-SP, Brasil. Os objetivos principais foram: (1) Examinar, em seis costões rochosos, séries temporais de assentamento larval de decápodos, afim de entender forçantes físicas associadas ao transporte em direção à costa, (2) Localizar agregações larvais no plâncton neustônico costeiro, para inferir a natureza das agregações e verificar a ocorrência de migração vertical em decápodos, (3) testar a relação entre suprimento e assentamento larval, a diferentes escalas espaciais. O papel da competência larval foi discutido à luz de evidências obtidas através amostragem planctônica em diferentes áreas dentro das enseadas e comparações inter e intra-taxonômicas. Entre maio e agosto de 2005, foram realizadas coletas, a cada dois dias, para estimar a taxa de assentamento larval de grupos de decápodes em coletores artificiais. Durante dois períodos de maior assentamento de Pachygrapsus transversus, a ocorrência nictemeral neustônica, desta e de outras espécies de decápodes foi avaliada. Sub-amostras de megalopas de brachyura foram separadas para mensuração do nível de competência larval, através do cultivo de larvas, obtendo estimativas do tempo decorrido até a muda, e através do estágio de muda de indivíduos previamente fixados. Modelos Log-lineares foram utilizados para testar o efeito do vento e da maré na taxa de assentamento, e os resultados foram confrontados com cenários de correntes superficiais fornecidos pelo modelo hidrodinâmico de circulação. Amostras planctônicas revelaram que agregações larvais são coincidentes com zonas de convergência, provavelmente causadas pela formação de ondas internas. Um conjunto de espécies de camarão sofre migração vertical típica, enquanto um padrão de migração reversa foi encontrado para grapsídeos, ix o que pode auxiliar no transporte em direção à costa para as áreas de assentamento devido a brisas marinhas. Portunídeos não apresentaram tendência de migração vertical. A competência parece ser melhor estimada através do método de cultivo. Em áreas dentro das enseadas, megalopas de portunídeos parecem ser mais competentes ao assentamento que as de grapsídeos, provavelmente por estarem mais próximas aos locais de assentamento. Megalopas neustônicas de P. transversus coletadas dentro das enseadas são menos competentes que as obtidas em coletores artificiais. O estágio transicional não foi capturado, sugerindo que a transição para o bentos e o desenvolvimento até estágios competentes são mais complexos do que normalmente assumido. Não foi encontrada correlação significativa entre o suprimento de megalopas e a taxa de assentamento. A taxa de assentamento de decápodes, que habitam a região entre-marés de costões rochosos, é aparentemente dependente da interação das forçantes de vento, maré e comportamento larval. As taxas de assentamento de grapsídeos e palemonídeos seguem o ritmo semi-lunar, com ocorrência máxima próximo às marés de quadratura, corroborando a hipótese de que ondas internas geradas pela maré são também responsáveis pelo transporte de decápodos costeiros. Ventos em direção à costa foram identificados como agentes chave, apesar de dependentes do regime de marés, enquanto que ventos de nordeste provavelmente trazem larvas de locais de produção. Padrões de circulação costeira e contrastes de assentamento consistentes espacialmente indicam que a Enseada do Flamengo é uma importante área de deposição de larvas. / The present study was carried out along a coastline including two bays in Ubatuba, SP, Brazil. The main objectives were (1) examine, at six different rocky shores, the time series of decapod larval settlement in order to understand the physical forcing associated to onshore transport, (2) locate neustonic plankton larval aggregations over the diel cycle to infer on the nature of patchiness and to verify the occurrence of vertical migration in the decapod species examined, and (3) test the relationship between larval supply and settlement, at different spatial scales. The role of larval competency was discussed based on evidence obtained by sampling the plankton at distinct within-bay areas and comparing the results both within and between taxa. From May to August 2005, samples were carried out every other day to estimate the larval settlement rate of two different decapod assemblages in artificial collectors. During two settlement pulses recorded for the megalopae of Pachygrapsus transversus, the nycthemeral neustonic occurrence of that and other decapod stages was assessed. Subsamples of brachyuran megalopae were separated to measure their competency level, by means of both larval rearing, thus obtaining time-to-moult estimates, and microscopic moult-staging of readily fixed individuals. Log-linear models were used to test the effect of wind and tide range on settlement rate, and results were confronted to possible scenarios of surface currents provided by hydrodynamic numerical modeling. Planktonic samples revealed that patches of larvae are coincident to slick fringes, probably caused by the formation of internal waves. A suite of shrimp species undertakes a typical diel vertical migration, while a reversed pattern was found for grapsids, which may assist the final onshore transport to settlement grounds due to marine breezes. Swimming crabs did not show any vertical xi migration trend. Competency is suggested to be better estimated using the larval rearing method. In within-bay areas, portunid megalopae appeared to be more competent to settle than those of grapsids, probably because the former are closer to settlement substrates than the latter, which should be first advected to the nearshore. Within-taxon comparisons revealed that neustonic megalopae of P. transversus collected in within-bay areas are less competent than those obtained from benthic collectors. A transitional stage was missing, suggesting that the transition to the benthos and the development into competent stages are more complex than usually assumed. Significant correlations between megalopae supply and settlement rate were not found. The settlement rate of decapods inhabiting the intertidal region of the rocky coast sampled is apparently dependent on the interaction of wind and tide forcing coupled with larval behaviour. Settlement rate of both grapsids and palaemonids follows a semilunar rhythm, with maxima occurring near neap tides, corroborating the hypothesis that tidally-driven internal motions are also responsible for onshore larval transport coastal decapods. Onshore wind-forcing was found to be the key agent, although dependent on the tide regime, while winds blowing from northeast are suggested to bring larvae from source sites. Coastal circulation patterns and spatially-consistent settlement contrasts indicate that Flamengo Bay is an important area of larval deposition.

by Fong Man Kim. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2001. / Includes bibliographical references (leaves 127-137). / Abstracts in English and Chinese. / Thesis committee --- p.i / Acknowledgements --- p.ii / Abstract --- p.iii / List of figures --- p.viii / List of tables --- p.xi / Abbreviations --- p.xii / Table of contents --- p.xiv / Chapter CHAPTER 1 --- GENERAL INTRODUCTION --- p.1 / Chapter CHAPTER 2 --- LITERATURE REVIEW --- p.3 / Chapter 2.1 --- MIH from Penaeus vannamei --- p.3 / Chapter 2.1.1 --- General Introduction to P. vannamei --- p.3 / Chapter 2.1.1.1 --- Morphology --- p.3 / Chapter 2.1.1.2 --- Geographical distribution --- p.5 / Chapter 2.1.1.3 --- Economic value --- p.5 / Chapter 2.1.2 --- Physiology of Molting in Crustacean --- p.7 / Chapter 2.1.2.1 --- The molt cycle --- p.7 / Chapter 2.1.2.2 --- Physiological effects of ecdysone --- p.8 / Chapter 2.1.2.3 --- Regulation of the secretion of ecdysone --- p.9 / Chapter 2.1.2.4 --- Physiological effects of Molt-inhibiting hormone --- p.10 / Chapter 2.1.3 --- Cloning of MIH cDNA from P. vannamei --- p.14 / Chapter 2.1.3.1 --- Molecular identity of MIH --- p.14 / Chapter 2.1.3.2 --- Cloning of MIH cDNA --- p.15 / Chapter 2.1.3.3 --- Comparison of the cloned MIH-like cDNA with the CHH/MIH/VIH peptide family --- p.16 / Chapter 2.2 --- Plants as Bioreactors --- p.20 / Chapter 2.2.1 --- Principles & Techniques --- p.20 / Chapter 2.2.2 --- Advantages of plant bioreactors --- p.21 / Chapter 2.2.3 --- Tobacco expression system --- p.22 / Chapter 2.2.3.1 --- Tobacco as model plants --- p.22 / Chapter 2.2.3.2 --- Transformation methods --- p.23 / Chapter 2.2.4 --- Phaseolin --- p.26 / Chapter CHAPTER 3 --- EXPRESSION OF MIH IN TRANSGENIC TOBACCO --- p.28 / Chapter 3.1 --- Introduction --- p.28 / Chapter 3.2 --- Materials & Methods --- p.29 / Chapter 3.2.1 --- Chemicals --- p.29 / Chapter 3.2.2 --- Plant materials --- p.29 / Chapter 3.2.3 --- Bacterial strains and plasmid vectors --- p.30 / Chapter 3.2.4 --- Construction of chimeric genes - --- p.30 / Chapter 3.2.4.1 --- PCR amplification of MIH --- p.30 / Chapter 3.2.4.2 --- Cloning of PCR-amplified MIH into vector pET --- p.31 / Chapter 3.2.4.3 --- Cloning of MIH into vector pBK/Phas-sp and pTZ/Phas --- p.31 / Chapter 3.2.4.4 --- Cloning of MIH into binary vector pBI121 --- p.32 / Chapter 3.2.5 --- Transformation of Agrobacterium with pBI121/Phas-sp-MIH and pBI121 /Phas-MIH by electroporation --- p.39 / Chapter 3.2.6 --- Transformation of tobacco --- p.40 / Chapter 3.2.7 --- Selection of transgenic plants --- p.41 / Chapter 3.2.8 --- GUS assay --- p.42 / Chapter 3.2.9 --- Extraction of leaf genomic DNA --- p.43 / Chapter 3.2.10 --- Extraction of total RNA from developing seeds --- p.44 / Chapter 3.2.11 --- Synthesis of DIG-labeled DNA and RNA probes --- p.45 / Chapter 3.2.12 --- Southern blot analysis of genomic DNA --- p.47 / Chapter 3.2.13 --- Reverse transcriptase - polymerase chain reaction (RT-PCR) --- p.47 / Chapter 3.2.14 --- Northern blot analysis of total RNA --- p.48 / Chapter 3.2.15 --- Protein extraction and tricine-SDS-PAGE --- p.49 / Chapter 3.2.16 --- Purification of 6xHis-tag proteins --- p.50 / Chapter 3.2.17 --- Western blot analysis --- p.50 / Chapter 3.2.18 --- In vitro transcription & translation --- p.52 / Chapter 3.2.18.1 --- Construction of transcription vector containing the chimeric MIH gene --- p.52 / Chapter 3.2.18.2 --- In vitro transcription --- p.56 / Chapter 3.2.18.3 --- In vitro translation --- p.56 / Chapter 3.2.19 --- Particle bombardment --- p.57 / Chapter 3.2.19.1 --- Construction of MIH-GUSN fusion chimeric genes --- p.57 / Chapter 3.2.19.2 --- Conditions of particle bombardment --- p.63 / Chapter 3.2.20 --- Codon modification of MIH gene --- p.63 / Chapter 3.3 --- Results --- p.73 / Chapter 3.3.1 --- Construction of chimeric MIH genes --- p.73 / Chapter 3.3.2 --- "Tobacco transformation, selection and regeneration" --- p.73 / Chapter 3.3.3 --- Detection of GUS activity --- p.74 / Chapter 3.3.4 --- Southern blot analysis --- p.79 / Chapter 3.3.5 --- Detection of MIH transcript in transgenic tobacco --- p.83 / Chapter 3.3.5.1 --- RT-PCR --- p.83 / Chapter 3.3.5.2 --- Northern blot analysis --- p.86 / Chapter 3.3.6 --- Detection of MIH protein by Tricine-SDS-PAGE --- p.86 / Chapter 3.3.7 --- Detection of MIH protein by western blot analysis --- p.88 / Chapter 3.3.7.1 --- Western blot analysis using Anti-MIH antibody --- p.88 / Chapter 3.3.7.2 --- Western blot analysis using Anti-His antibody --- p.90 / Chapter 3.3.7.3 --- Western blot analysis using Anti-MIHA & Anti-MIHB antibodies --- p.90 / Chapter 3.3.8 --- Purification of 6xHis-tag proteins by Ni-NTA column --- p.94 / Chapter 3.3.8.1 --- Western blot analysis of proteins purified by Ni-NTA column --- p.97 / Chapter 3.3.9 --- In vitro transcription and translation --- p.100 / Chapter 3.3.9.1 --- In vitro transcription --- p.100 / Chapter 3.3.9.2 --- In vitro translation --- p.100 / Chapter 3.3.10 --- Particle bombardments --- p.103 / Chapter 3.3.10.1 --- Transient expression of MIH in soybean & tobacco leaves --- p.103 / Chapter CHAPTER 4 --- DISCUSSION --- p.107 / Chapter 4.1 --- Transient expression of MIH genes --- p.109 / Chapter 4.1.1 --- In vitro transcription and translation --- p.109 / Chapter 4.1.2 --- Particle bombardments --- p.220 / Chapter 4.2 --- Post-transcriptional gene silencing (PTGS) --- p.114 / Chapter 4.2.1 --- Post-transcriptional cis-inactivation --- p.114 / Chapter 4.2.2 --- Post-transcriptional trans-inactivation --- p.116 / Chapter 4.2.3 --- MIH gene and PTGS --- p.118 / Chapter 4.3 --- Codon usage --- p.119 / Chapter 4.3.1 --- Codon usage of MIH in plants --- p.120 / Chapter 4.3.2 --- Codon modification of MIH and further study on MIH expression in plants --- p.122 / Chapter 4.4 --- Post-translational protein degradation --- p.123 / Chapter 4.4.1 --- Construction of LRP-MIH fusion proteins --- p.123 / CONCLUSION --- p.125 / REFERENCES --- p.127

Made available in DSpace on 2015-04-17T14:55:26Z (GMT). No. of bitstreams: 1 arquivototal.pdf: 1732626 bytes, checksum: 1ea01eff6b48d045cb5829753b4efd89 (MD5) Previous issue date: 2013-01-25 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / The Order Cladocera is amongst the most studied scientific groups on continental aquatic environments, given that they provide valuable information regarding energy transfer between trophic levels. Cladocerans play important ecological roles by serving as food for higher predators (invertebrates and vertebrates) and by regulating the biomass of freshwater phytoplankton. This study aimed to evaluate the influence of vertebrate predators on Ceriodaphnia cornuta using laboratory experiments. The influence of potential predators on C. cornuta was experimentally tested based on density comparisons along five days (2-day interval between analyses) in the presence of two vertebrate predators, namely, the Nile Tilapia (Oreochromis niloticus, Cichlidae) and the Twospot astyanax (Astyanax bimaculatus,Characidae). C. cornuta individuals remained permanently isolated from predators by a 45 μm mesh size net during the experiment and, therefore, predation rates were not accounted for throughout the study. Furthermore, fertility rate and size were also evaluated alongside with density of C. cornuta and environmental variables sampled included water temperature, pH and dissolved oxygen. Results revealed a higher adult mortality on the former experimental days (due to acclimation or age). Further C. cornuta density increased on both predator treatments (O. niloticus and A. bimaculatus), but was fairly higher on the control treatment (no predators; 5th day). Fertility rate increased on the last experiment days on both predator treatments. Higher body size values were also observed on the last experiment days of the control treatment. Results of the present study suggest that, in the absence of vertebrate predators, C. cornuta showed a somewhat higher population growth and larger body size. / A Ordem Cladocera é uma das mais estudadas e importantes nos ambientes aquáticos continentais, pois, além de fornecerem muitas informações sobre a transferência de energia ao longo da cadeia trófica, algumas de suas espécies apresentam uma grande importância no controle do fitoplâncton da água doce e ao mesmo tempo, algumas espécies são presas preferidas de vertebrados e invertebrados. O trabalho teve como foco avaliar a influência de predadores vertebrados sobre a história de vida da espécie Ceriodaphnia cornuta,através de experimentos realizados em laboratório. A influência de potenciais predadores sobre a C. cornutafoi avaliada experimentalmente, a partir das comparações das densidades do cladócero, ao longo de cinco dias, com análises a cada dois dias, na presença de dois predadores vertebrados, tilápia (Oreochromis niloticus, Cichlidae) e piaba (Astyanax bimaculatus,Characidae). Os indivíduos de C. cornuta ficaram separados dos predadores, por uma tela de 45 μm, logo as interferências que foram analisadas excluíram a taxa de predação. Foram analisados para além das densidades, as taxas de fecundidade e o comprimento dos organismos. Alguns parâmetros ambientais aquáticos dos experimentos também foram analisados (temperatura, pH, oxigênio dissolvido). Os principais resultados deste trabalho incluem maior mortalidade dos adultos nos primeiros dias (aclimatação ou idade),um aumento na densidade de C. cornuta foi observado em todos os tratamentos (com piaba e com tilápia), com destaque para o quinto dia nos aquários controle, ou seja, C. cornuta, sem a presença de predadores. Verificou-seo aumento da fecundidade nos últimos dias de avaliaçãonos aquários com tilápia e piaba. Nos dados referentes ao comprimento do corpo, verificou-se que nos últimos dias de experimento o comprimento dos indivíduos foi maior no controle, apesar de não haver dierenças estatísticas.Os resultados obtidos com este experimento mostraram que na ausência de um predador vertebrado, C. cornuta apresentou um crescimento populacional maior e um maior comprimento do corpo.

O presente estudo foi realizado ao longo de uma linha de costa, incluindo duas enseadas em Ubatuba-SP, Brasil. Os objetivos principais foram: (1) Examinar, em seis costões rochosos, séries temporais de assentamento larval de decápodos, afim de entender forçantes físicas associadas ao transporte em direção à costa, (2) Localizar agregações larvais no plâncton neustônico costeiro, para inferir a natureza das agregações e verificar a ocorrência de migração vertical em decápodos, (3) testar a relação entre suprimento e assentamento larval, a diferentes escalas espaciais. O papel da competência larval foi discutido à luz de evidências obtidas através amostragem planctônica em diferentes áreas dentro das enseadas e comparações inter e intra-taxonômicas. Entre maio e agosto de 2005, foram realizadas coletas, a cada dois dias, para estimar a taxa de assentamento larval de grupos de decápodes em coletores artificiais. Durante dois períodos de maior assentamento de Pachygrapsus transversus, a ocorrência nictemeral neustônica, desta e de outras espécies de decápodes foi avaliada. Sub-amostras de megalopas de brachyura foram separadas para mensuração do nível de competência larval, através do cultivo de larvas, obtendo estimativas do tempo decorrido até a muda, e através do estágio de muda de indivíduos previamente fixados. Modelos Log-lineares foram utilizados para testar o efeito do vento e da maré na taxa de assentamento, e os resultados foram confrontados com cenários de correntes superficiais fornecidos pelo modelo hidrodinâmico de circulação. Amostras planctônicas revelaram que agregações larvais são coincidentes com zonas de convergência, provavelmente causadas pela formação de ondas internas. Um conjunto de espécies de camarão sofre migração vertical típica, enquanto um padrão de migração reversa foi encontrado para grapsídeos, ix o que pode auxiliar no transporte em direção à costa para as áreas de assentamento devido a brisas marinhas. Portunídeos não apresentaram tendência de migração vertical. A competência parece ser melhor estimada através do método de cultivo. Em áreas dentro das enseadas, megalopas de portunídeos parecem ser mais competentes ao assentamento que as de grapsídeos, provavelmente por estarem mais próximas aos locais de assentamento. Megalopas neustônicas de P. transversus coletadas dentro das enseadas são menos competentes que as obtidas em coletores artificiais. O estágio transicional não foi capturado, sugerindo que a transição para o bentos e o desenvolvimento até estágios competentes são mais complexos do que normalmente assumido. Não foi encontrada correlação significativa entre o suprimento de megalopas e a taxa de assentamento. A taxa de assentamento de decápodes, que habitam a região entre-marés de costões rochosos, é aparentemente dependente da interação das forçantes de vento, maré e comportamento larval. As taxas de assentamento de grapsídeos e palemonídeos seguem o ritmo semi-lunar, com ocorrência máxima próximo às marés de quadratura, corroborando a hipótese de que ondas internas geradas pela maré são também responsáveis pelo transporte de decápodos costeiros. Ventos em direção à costa foram identificados como agentes chave, apesar de dependentes do regime de marés, enquanto que ventos de nordeste provavelmente trazem larvas de locais de produção. Padrões de circulação costeira e contrastes de assentamento consistentes espacialmente indicam que a Enseada do Flamengo é uma importante área de deposição de larvas. / The present study was carried out along a coastline including two bays in Ubatuba, SP, Brazil. The main objectives were (1) examine, at six different rocky shores, the time series of decapod larval settlement in order to understand the physical forcing associated to onshore transport, (2) locate neustonic plankton larval aggregations over the diel cycle to infer on the nature of patchiness and to verify the occurrence of vertical migration in the decapod species examined, and (3) test the relationship between larval supply and settlement, at different spatial scales. The role of larval competency was discussed based on evidence obtained by sampling the plankton at distinct within-bay areas and comparing the results both within and between taxa. From May to August 2005, samples were carried out every other day to estimate the larval settlement rate of two different decapod assemblages in artificial collectors. During two settlement pulses recorded for the megalopae of Pachygrapsus transversus, the nycthemeral neustonic occurrence of that and other decapod stages was assessed. Subsamples of brachyuran megalopae were separated to measure their competency level, by means of both larval rearing, thus obtaining time-to-moult estimates, and microscopic moult-staging of readily fixed individuals. Log-linear models were used to test the effect of wind and tide range on settlement rate, and results were confronted to possible scenarios of surface currents provided by hydrodynamic numerical modeling. Planktonic samples revealed that patches of larvae are coincident to slick fringes, probably caused by the formation of internal waves. A suite of shrimp species undertakes a typical diel vertical migration, while a reversed pattern was found for grapsids, which may assist the final onshore transport to settlement grounds due to marine breezes. Swimming crabs did not show any vertical xi migration trend. Competency is suggested to be better estimated using the larval rearing method. In within-bay areas, portunid megalopae appeared to be more competent to settle than those of grapsids, probably because the former are closer to settlement substrates than the latter, which should be first advected to the nearshore. Within-taxon comparisons revealed that neustonic megalopae of P. transversus collected in within-bay areas are less competent than those obtained from benthic collectors. A transitional stage was missing, suggesting that the transition to the benthos and the development into competent stages are more complex than usually assumed. Significant correlations between megalopae supply and settlement rate were not found. The settlement rate of decapods inhabiting the intertidal region of the rocky coast sampled is apparently dependent on the interaction of wind and tide forcing coupled with larval behaviour. Settlement rate of both grapsids and palaemonids follows a semilunar rhythm, with maxima occurring near neap tides, corroborating the hypothesis that tidally-driven internal motions are also responsible for onshore larval transport coastal decapods. Onshore wind-forcing was found to be the key agent, although dependent on the tide regime, while winds blowing from northeast are suggested to bring larvae from source sites. Coastal circulation patterns and spatially-consistent settlement contrasts indicate that Flamengo Bay is an important area of larval deposition.