Made available in DSpace on 2016-08-31T13:19:26Z (GMT). No. of bitstreams: 1 SuzanyILM_DISSERT.pdf: 1345227 bytes, checksum: 8df554393ca0a302d49e17dbe4828ca0 (MD5) Previous issue date: 2015-04-13 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Prochilodus brevis (curimatã) is a migratory and endemic fish species of the hydrographic basin in semiarid region, occurring in rivers and reservoirs. The aims of this study was evaluate the population structure species in two reservoirs (Pau dos Ferros and Santa Cruz Reservoirs) that are located in the hydrographic basin of the Apodi/Mossoró River. The biological material was obtained through quarterly collections carried out from February/2011 to November/2012, using gillnets from 12 to 70mm. We evaluated Catch Per Unit Effort by the number (CPUEn), the length structure in 3 cm classes, sex ratio, the length-weight relationship (Wt), condition factor (K), the length at first maturity and the gonadosomatic index (GSI). The Pau dos Ferros Reservoir showed a higher abundance and better structured population with capture young individuals and adults in higher abundance. In the Pau dos Ferros Reservoir the distribution of the species was homogeneous, while in the Santa Cruz reservoir the species had a higher capture at at fluvial region. The sex ratio remained within the expected of 1:1 both the Pau dos Ferros reservoir as to the Santa Cruz reservoir. Isometric growth of the species (b=3.0109) was in Pau dos Ferros reservoir and allometric negative (b=2.6710) in the Santa Cruz Reservoir. Furthermore, the condition factor indicated that the population of the species in Pau dos Ferros reservoir had a better physiological condition. The length at first maturity for the Pau dos Ferros reservoir and Santa Cruz were 16.97cm and 17.40cm, respectively. Reproductive investment was similar between the reservoirs; however, for the population of the Santa Cruz reservoir is not being reflected in the recruitment of new individuals to the population, since that capture of juveniles was low. The results found suggested that the species maintain viable populations in Pau dos Ferros Reservoir, on the other hand in the Santa Cruz reservoir, probably in this reservoir species not found appropriate places to spawn and/or development of juveniles. Thus, the results suggested that the construction of the Santa Cruz reservoir was detrimental to the population of P. brevis and if appropriate measures are not taken handling species may disappear from the reservoir / O Prochilodus brevis (curimatã) é uma espécie migradora, endêmicas das bacias hidrográficas da região do semiárido, com ocorrência em rios e reservatórios da região. O presente trabalho teve como objetivo avaliar a estrutura populacional da espécie em dois reservatórios (Pau dos Ferros e Santa Cruz) localizados na bacia hidrográfica do rio Apodi/Mossoró. O material biológico foi obtido através de coletas trimestrais realizadas de fevereiro de 2011 a novembro de 2012, usando rede de espera. Foram analisados a Captura Por Unidade de Esforço (CPUE), a estrutura em comprimento em classes de 3 cm, a proporção sexual, a relação comprimento-peso (Wt), o fator de condição (K), o comprimento de primeira maturação e o índice gonadossomático (IGS). O reservatório de Pau dos Ferros apresentou uma maior abundância e uma população melhor estruturada com captura maior de indivíduos jovens e adultos. Diferente do reservatório de Pau dos Ferros em que a distribuição da espécie foi homogênea no reservatório de Santa Cruz a espécie teve uma maior captura nos pontos mais próximos ao rio. A proporção sexual permaneceu dentro do esperado DE 1:1 tanto para o reservatório de Pau dos Ferros como para o reservatório de Santa Cruz. A espécie apresentou crescimento isométrico (b=3,0109) no reservatório de Pau dos Ferros e alométrico negativo (b=2,6710) no reservatório de Santa Cruz. Além disso, o fator de condição indicou que a população da espécie no reservatório de Pau dos Ferros apresenta um estado fisiológico melhor. O comprimento de primeira maturação para o reservatório de Pau dos Ferros e Santa Cruz foi de 16,97cm e 17,40cm, respectivamente. O investimento reprodutivo foi semelhante entre os reservatórios, no entanto, para a população do reservatório de Santa Cruz não está se refletindo no recrutamento de novos indivíduos para a população, uma vez que, a captura de juvenis foi baixa nesse reservatório. Através desses resultados constatou-se que a espécie mantem populações viáveis no reservatório de Pau dos Ferros por outro lado no reservatório de Santa Cruz, provavelmente a espécie não encontrou locais apropriados para desovar e/ou desenvolvimento dos juvenis. Assim, os resultados sugerem que a construção do reservatório de Santa Cruz foi prejudicial para população de P. brevis e caso medidas adequadas de manejo não sejam tomadas a espécie poderá desaparecer do reservatório

Coral cover is in decline on a global scale, with increased mortality events being attributed to a number of global and regional stressors. While the impacts of global stressors (e.g. sea surface temperature rise, ocean acidification) are well documented, there is growing interest in identifying and understanding the impacts of regional stressors. The reason for this change in focus is that regional stressors can often work in combination, sometimes synergistically, with global stressors and that stressors on a regional scale tend to be more easily mitigated by management practices. One regional stressor that impacts a myriad of marine organisms in the southeastern United States is the annual red tide blooms produced by the dinoflagellate, Karenia brevis. Their impacts, along with the lipid soluble polyether neurotoxins they produce, termed brevetoxins, are well studied in economically important organisms, such as bivalves. However, little is known of their impacts on organisms that possess ecological importance, such as species of scleractinian coral. To address this gap in knowledge, a multifaceted study is discussed herein, which evaluated the effects of ecologically relevant concentrations of K. brevis and associated brevetoxins on different coral life history stages and coral species. The second chapter addresses the impacts of red tide on larval behavior, settlement and survival of the coral species Porites astreoides, as well as impacts of photochemical efficiency and oxidative stress within different coral species (P. astreoides larvae, P. astreoides adults, Acropora cervicornis, Cladocora arbuscula, and Phyllangia americana). The third chapter confers the use of broad-scale proteomic analysis to identify the cellular response of the non-model coral species, P. astreoides, following exposure to red tide. Coral larvae actively avoided both medium and high bloom conditions of K. brevis and brevetoxins, while percent larval settlement and survival were impacted following exposure to high bloom concentrations of K. brevis. Photochemical efficiency of in hospite Symbiodinium was reduced following exposure to both K. brevis and brevetoxin in P. astreoides larvae, as well as exposure to K. brevis in P. astreoides adults, while being unimpacted in A. cervicornis. Compared to controls, high bloom conditions resulted in an increase in biomarkers of lipid peroxidation in C. arbuscula. This was also seen in P. astreoides larvae at 24 hours; however, this difference was indistinguishable following 48 hours. Surprisingly, no other biomarker of oxidative stress analyzed were impacted. Broad-scale proteomic analysis of P. astreoides following exposure to red tide conditions revealed variable changes in proteome expression depending on if the corals were exposed to K. brevis or brevetoxins. Exposure to brevetoxins resulted in differential expression of proteins related to DNA organization, chromatin formation and transcription expression; while exposure to K. brevis resulted in differential expression of proteins related to redox homeostasis, protein folding, energy metabolism, and production of reactive oxygen species. The results of this study demonstrate the potential for annual red tide blooms to act as a regional stressor on coral species. They highlight the ability of red tide conditions to negatively impact coral at multiple life history stages and that the extent of these effects may be species specific. They also provide further incite of coral’s response to red tide exposure at the cellular level.

Metabolomics is the methodical scientific study of biochemical processes associated with the metabolome—which comprises the entire collection of metabolites in any biological entity. Metabolome changes occur as a result of modifications in the genome and proteome, and are, therefore, directly related to cellular phenotype. Thus, metabolomic analysis is capable of providing a snapshot of cellular physiology. Untargeted metabolomics is an impartial, all-inclusive approach for detecting as many metabolites as possible without a priori knowledge of their identity. Hence, it is a valuable exploratory tool capable of providing extensive chemical information for discovery and hypothesis-generation regarding biochemical processes. A history of metabolomics and advances in the field corresponding to improved analytical technologies are described in Chapter 1 of this dissertation. Additionally, Chapter 1 introduces the analytical workflows involved in untargeted metabolomics research to provide a foundation for Chapters 2 – 5. Part I of this dissertation which encompasses Chapters 2 – 3 describes the utilization of mass spectrometry (MS)-based untargeted metabolomic analysis to acquire new insight into cancer detection. There is a knowledge deficit regarding the biochemical processes of the origin and proliferative molecular mechanisms of many types of cancer which has also led to a shortage of sensitive and specific biomarkers. Chapter 2 describes the development of an in vitro diagnostic multivariate index assay (IVDMIA) for prostate cancer (PCa) prediction based on ultra performance liquid chromatography-mass spectrometry (UPLC-MS) metabolic profiling of blood serum samples from 64 PCa patients and 50 healthy individuals. A panel of 40 metabolic spectral features was found to be differential with 92.1% sensitivity, 94.3% specificity, and 93.0% accuracy. The performance of the IVDMIA was higher than the prevalent prostate-specific antigen blood test, thus, highlighting that a combination of multiple discriminant features yields higher predictive power for PCa detection than the univariate analysis of a single marker. Chapter 3 describes two approaches that were taken to investigate metabolic patterns for early detection of ovarian cancer (OC). First, Dicer-Pten double knockout (DKO) mice that phenocopy many of the features of metastatic high-grade serous carcinoma (HGSC) observed in women were studied. Using UPLC-MS, serum samples from 14 early-stage tumor DKO mice and 11 controls were analyzed. Iterative multivariate classification selected 18 metabolites that, when considered as a panel, yielded 100% accuracy, sensitivity, and specificity for early-stage HGSC detection. In the second approach, serum metabolic phenotypes of an early-stage OC pilot patient cohort were characterized. Serum samples were collected from 24 early-stage OC patients and 40 healthy women, and subsequently analyzed using UPLC-MS. Multivariate statistical analysis employing support vector machine learning methods and recursive feature elimination selected a panel of metabolites that differentiated between age-matched samples with 100% cross-validated accuracy, sensitivity, and specificity. This small pilot study demonstrated that metabolic phenotypes may be useful for detecting early-stage OC and, thus, supports conducting larger, more comprehensive studies. Many challenges exist in the field of untargeted metabolomics. Part II of this dissertation which encompasses Chapters 4 – 5 focuses on two specific challenges. While metabolomic data may be used to generate hypothesis concerning biological processes, determining causal relationships within metabolic networks with only metabolomic data is impractical. Proteins play major roles in these networks; therefore, pairing metabolomic information with that acquired from proteomics gives a more comprehensive snapshot of perturbations to metabolic pathways. Chapter 4 describes the integration of MS- and NMR-based metabolomics with proteomics analyses to investigate the role of chemically mediated ecological interactions between Karenia brevis and two diatom competitors, Asterionellopsis glacialis and Thalassiosira pseudonana. This integrated systems biology approach showed that K. brevis allelopathy distinctively perturbed the metabolisms of these two competitors. A. glacialis had a more robust metabolic response to K. brevis allelopathy which may be a result of its repeated exposure to K. brevis blooms in the Gulf of Mexico. However, K. brevis allelopathy disrupted energy metabolism and obstructed cellular protection mechanisms including altering cell membrane components, inhibiting osmoregulation, and increasing oxidative stress in T. pseudonana. This work represents the first instance of metabolites and proteins measured simultaneously to understand the effects of allelopathy or in fact any form of competition. Chromatography is traditionally coupled to MS for untargeted metabolomics studies. While coupling chromatography to MS greatly enhances metabolome analysis due to the orthogonality of the techniques, the lengthy analysis times pose challenges for large metabolomics studies. Consequently, there is still a need for developing higher throughput MS approaches. A rapid metabolic fingerprinting method that utilizes a new transmission mode direct analysis in real time (TM-DART) ambient sampling technique is presented in Chapter 5. The optimization of TM-DART parameters directly affecting metabolite desorption and ionization, such as sample position and ionizing gas desorption temperature, was critical in achieving high sensitivity and detecting a broad mass range of metabolites. In terms of reproducibility, TM-DART compared favorably with traditional probe mode DART analysis, with coefficients of variation as low as 16%. TM-DART MS proved to be a powerful analytical technique for rapid metabolome analysis of human blood sera and was adapted for exhaled breath condensate (EBC) analysis. To determine the feasibility of utilizing TM-DART for metabolomics investigations, TM-DART was interfaced with traveling wave ion mobility spectrometry (TWIMS) time-of-flight (TOF) MS for the analysis of EBC samples from cystic fibrosis patients and healthy controls. TM-DART-TWIMS-TOF MS was able to successfully detect cystic fibrosis in this small sample cohort, thereby, demonstrating it can be employed for probing metabolome changes. Finally, in Chapter 6, a perspective on the presented work is provided along with goals on which future studies may focus.