Photosynthetic Microbial Fuel Cell (p-MFC) research aims to develop devices containing photosynthetic micro-organisms to produce electricity. Micro-organisms within the device photosynthesise carbohydrates under illumination, and produce reductive equivalents (excess electrons) from both carbohydrate production and the subsequent carbohydrate break down. Redox mediators are utilised to shuttle electrons between the organism and the electrode. The mediator is reduced by the micro-organism and subsequently re-oxidised at the electrode. However this technology is in its early stages and extensive research is required for p-MFC devices to become economically viable. A basic p-MFC device containing a potassium ferricyanide mediator and the algae Chlorella vulgaris was assembled and tested. From these initial experiments it was realised that much more work was required to characterise cell and redox mediator activities occurring within the device. There is very little p-MFC literature dealing with cellular interaction with redox mediators, but without this knowledge the output of complete p-MFC devices can not be fully understood. This thesis presents research into the reduction of redox mediators by the micro-organisms, including rates of mediator reduction and factors affecting the rate. Both electrochemical and non-electrochemical techniques are used and results compared. Additionally, cellular effects relating to the presence of the mediator are studied; crucial to provide limits within which p-MFCs must be used. After basic characterisation, this thesis presents work into the optimisation of the basic p-MFC. Different redox mediators, photosynthetic species and anodic materials are investigated. Importantly, it is only through fundamental characterization to improve understanding that p-MFCs can be optimised.

abstract: Utilizing both 16S and 18S rRNA sequencing alongside energetic calculations from geochemical measurements offers a bridged perspective of prokaryotic and eukaryotic community diversities and their relationships to geochemical diversity. Yellowstone National Park hot spring outflows from varied geochemical compositions, ranging in pH from < 2 to > 9 and in temperature from < 30°C to > 90°C, were sampled across the photosynthetic fringe, a transition in these outflows from exclusively chemosynthetic microbial communities to those that include photosynthesis. Illumina sequencing was performed to document the diversity of both prokaryotes and eukaryotes above, at, and below the photosynthetic fringe of twelve hot spring systems. Additionally, field measurements of dissolved oxygen, ferrous iron, and total sulfide were combined with laboratory analyses of sulfate, nitrate, total ammonium, dissolved inorganic carbon, dissolved methane, dissolved hydrogen, and dissolved carbon monoxide were used to calculate the available energy from 58 potential metabolisms. Results were ranked to identify those that yield the most energy according to the geochemical conditions of each system. Of the 46 samples taken across twelve systems, all showed the greatest energy yields using oxygen as the main electron acceptor, followed by nitrate. On the other hand, ammonium or ammonia, depending on pH, showed the greatest energy yields as an electron donor, followed by H2S or HS-. While some sequenced taxa reflect potential biotic participants in the sulfur cycle of these hot spring systems, many sample locations that yield the most energy from ammonium/ammonia oxidation have low relative abundances of known ammonium/ammonia oxidizers, indicating potentially untapped sources of chemotrophic energy or perhaps poorly understood metabolic capabilities of cultured chemotrophs. / Dissertation/Thesis / Masters Thesis Geological Sciences 2018

O objetivo desse trabalho foi avaliar os efeitos da radiação UV-B em indivíduos de linhagens de cultivados em laboratório. Foram utilizadas linhagens de gametófitos femininos de coloração vermelha (VMsubES/sub e VMSUBCE/SUB), verde (VD subCE/sub) e marrom-esverdeada (ME subCE/sub), derivados de populações dos Estados do Espírito Santo e Ceará. Dois experimentos distintos foram delineados: exposição à radiação UV-B por 5 horas (curto prazo - 2,01 W.msup-2/sup) e exposição por 3 horas diárias, durante 28 dias (longo prazo - 0,08 W.m-2). Ambos incluíram uma condição controle (sem radiação UV-B). As condições gerais de cultivo foram: água do mar enriquecida com von Stosch à 12,5% sem nitrato; nitrato (0,250mM); 25&#177;1supo/supC; 14L:10E; 70±10µmol.fótons.msup-2/sup.ssup-1/sup; sem aeração. No experimento de curta duração, foram avaliados os seguintes parâmetros: fotossíntese, composição pigmentar e ultra-estrutura. No experimento de longa duração, foram avaliados: crescimento, composição pigmentar e ultra-estrutura. Curvatura acentuada dos ápices após a exposição prolongada à UV-B e mudanças na coloração foram observadas principalmente em indivíduos da linhagem VMsubES/sub, cujos ápices adquiriram aspecto espiralado, quando cultivados em UV-B. Esses mesmos indivíduos apresentaram menores taxas de crescimento (TCs), quando comparados à linhagem selvagem do Estado do Ceará (VMSUBCE/SUB), sugerindo uma maior sensibilidade à radiação UV-B. Após exposição de 5 horas à UV-B, todas as linhagens apresentaram reduções no rendimento quântico efetivo (&nabla;F/Fm), com exceção de VMSUBCE/SUB, que não apresentou variações nos valores de &nabla;F/Fm no decorrer do experimento. As linhagens VMsubES/sub, VDSUBCE/SUBe MESUBCE/SUB apresentaram redução nesse parâmetro logo após a primeira hora de exposição à UV-B. Entretanto, esses valores mantiveram-se constantes para a linhagem MESUBCE/SUB ao longo do período experimental. Para a linhagem VDCE, verificou-se uma recuperação de F/Fm nos dois dias subseqüentes, o que sugere a ação de uma fotoinibição dinâmica, ao contrário das linhagens VMsubES/sub e MEsubCE/sub, cuja demora na recuperação caracteriza uma fotoinibição crônica. A linhagem VDSUBCE/SUB apresentou também maiores valores para a saturação luminosa (IK) após a exposição à UV-B, quando comparada às demais linhagens provenientes da população do Estado do Ceará (VMSUBCE/SUB e MEsubCE/sub). Na ausência de UV-B, as linhagens VMSUBCE/SUB e VMES apresentaram maiores valores para a eficiência fotossintetizante (&#945;ETR), quando comparadas às variantes cromáticas VDSUBCE/SUB e MEsubCE/sub. Entretanto, após a exposição à UV-B, a linhagem VMSUBCE/SUB apresentou maiores valores de ETR, quando comparada à linhagem VMsubES/sub, porém semelhantes aos observados nas linhagens VDSUBCE/SUB e MEsubCE/sub. Em curto prazo, não foram verificadas alterações nas concentrações de ficoeritrina (FE), ficocianina (FC) e aloficocianina nas diferentes linhagens, quando submetidas à radiação UV-B. Entretanto, um aumento na razão FE/FC foi observado para a linhagem VDsubCE/sub, sugerindo um aumento na proporção de FE, quando comparada à FC. Porém, reduções nas concentrações de ficoeritrina (FE), ficocianina e aloficocianina (AFC) foram observadas para indivíduos da linhagem VMSUBCE/SUB, após a exposição prolongada à UV-B, sugerindo uma possível utilização dessas substâncias no metabolismo celular, uma vez que as ficobiliproteínas, principalmente a FE, atuam como reserva de nitrogênio. Entretanto, as demais linhagens cultivadas em UV-B, apresentaram valores semelhantes de ficobiliproteínas aos verificados no controle. Observou-se um aumento na proporção FE/AFC na linhagem MEsubCE/sub, quando exposta à UV-B por 28 dias, sugerindo um aumento na porcentagem de FE em relação à AFC. Esse provável aumento nas concentrações de FE, em decorrência de sua síntese e reposição, favoreceria a aclimatação dos ficobilissomos a mudanças na irradiância. Concentrações semelhantes de clorofila a e carotenóides foram verificadas para as diferentes linhagens após a exposição à UV-B, independente do tempo de exposição, indicando a ausência de danos ao aparato fotossintetizante e a disponibilidade de pigmentos para o transporte de elétrons. Alterações ultraestruturais ocorreram em todas as linhagens cultivadas em UV-B em ambas condições experimentais. Os efeitos ultraestruturais foram observados principalmente nos cloroplastos, que apresentaram afastamento de tilacóides adjacentes, além da formação de vacúolos internos. Algumas células de indivíduos das linhagens VMSUBCE/SUB (longo prazo) e VDSUBCE/SUB (curto prazo) encontravam-se em avançado processo degenerativo, impossibilitando a visualização de membranas organelares. Observou-se também um maior número de grãos de amido nas linhagens VMsubES/sub e VDSUBCE/SUB cultivadas por 28 dias em UV-B, quando comparadas aos controles. Esse acúmulo de reserva poderia ser interpretado como uma incapacidade em metabolizar essas substâncias para atividades de síntese e reparo, necessárias para a manutenção do indivíduo quando em condições adversas, no caso a exposição à UV-B. Embora a linhagem VMsubES/subVMSUBCE/SUB tenha apresentado uma redução nas ficobiliproteínas, ela apresentou maiores TCs e menores alterações morfológicas. A linhagem MECE, embora tenha sofrido uma redução nas TCs, quando cultivada em UV-B, como as linhagens VDSUBCE/SUB e VMES, estas TCs mantiveram-se constantes ao longo do experimento, enquanto que nas demais linhagens elas diminuíram. A análise geral dos parâmetros avaliados sugere que as linhagens MESUBCE/SUB e VMCEVMSUBCE/SUB estariam aptas a lidar com um possível aumento nos níveis de UV-B incidente. Entretanto, diferenças na composição pigmentar observadas para a linhagem VDSUBCE/SUBsugerem uma menor sensibilidade desta a curto prazo, quando submetida a essa radiação. / não consta

Um novo conceito agro tecnológico em cultivo protegido foi a utilização de malhas coloridas, que permitem combinar a proteção física com a seleção de determinadas faixas do espectro da radiação solar para promover respostas fisiológicas desejadas reguladas pela luz. Dessa forma, o presente trabalho teve como objetivo avaliar a influência de malhas coloridas, azul, vermelha e preta (controle), no desenvolvimento de duas espécies de helicônias cultivadas em vaso: Heliconia H-01 e Helicônia ortrotricha Eclipse Total. Foram analisadas, para ambas as espécies, características agronômicas como altura e número de folhas finais da planta, quantidade de brotos; quantidade de estômatos por milímetro quadrado e análise espectral de folhas. A Helicônia H-01 não apresentou diferença significativa em relação ao número de folhas finais nos três tratamentos, porém as plantas cultivadas sob telado azul e vermelho apresentaram maiores alturas em relação ao controle. O telado azul apresentou maior número de brotos totais, mas não diferiu estatisticamente do controle no número de brotos lançados durante o tempo, sendo que o telado vermelho apresentou plantas com menor número de brotos. A maior quantidade de estômatos por mm2 foi encontrada nas plantas do telado vermelho, onde azul e controle não diferiram entre si. A análise de reflectância mostrou diferenças significativas na região visível, faixa de absorção da luz incidente pelos pigmentos fotossintetizantes. A H. ortrotricha não apresentou diferença significativa em relação ao número final de folhas, mas no telado azul apresentou a maior altura, seguido do vermelho e depois do controle. Não houve diferença significativa na quantidade de brotos lançado ao longo do tempo, mas o telado vermelho apresentou maior quantidade de brotos finais. Os tratamentos não apresentaram diferença significativa na quantidade de estômatos por mm2. Houve diferença significativa na curva espectral das folhas, principalmente na região do visível. / A new agro technology concept in greenhouse was the use of colored shade nets, which allow to combine the physical protection with the selection of spectral wavelengths of solar radiation to promote physiological responses regulated by light. This study aimed to evaluate the influence of colored shade nets, blue, red and black, in the development of two species of heliconias grown in pots: Heliconia H-01 and Heliconia ortrotricha Total Eclipse. The agronomic characteristics were studied for both species as plants height and number of leaves at the end of the experiment; number of shoots; number of stomata per square millimeter and spectral reflectance of leaves. There were no differences on the number of leaves between the treatments, but blue and red nets exhibited higher plants. Plants of blue net showed a greater number of shoots, and there were not statistically different from black net in the number of shoots during the study, and the red net plants showed fewer shoots than the other colored shade nets. The greater number of stomata per square millimeter was found in plants grown in red net, and blue and black nets did not showed significant differences. The spectral reflectance of H. H01 leaves showed significant differences in the absorption of visible wavelengths by the photosynthetic pigments. H. ortrotricha showed no significant difference on the number of leaves, but the plants of the blue net were highest and black net the lowest. There was no significant difference on the number of new shoots during the study, but the red net exhibited a greater number of shoots in the end of the study. All the treatments showed no significant difference in the number of stomata per square millimeter. There were significant differences in the spectral reflectance of leaves, mainly in the visible wavelengths.

abstract: Microscopic algae have been investigated extensively by researchers for decades for their ability to bioremediate wastewater and flue gas while producing valuable biomass for use as feed, fuel, fertilizer, nutraceutical, and other specialty products. Reports of the exciting commercial potential of this diverse group of organisms started appearing in the literature as early as the 1940’s. However, nearly 80 years later, relatively few successful commercial microalgae installations exist and algae have not yet reached agricultural commodity status. This dissertation examines three major bottlenecks to commercial microalgae production including lack of an efficient and economical cultivation strategy, poor management of volatile waste nutrients, and costly harvesting and post processing strategies. A chapter is devoted to each of these three areas to gain a better understanding of each bottleneck as well as strategies for overcoming them. The first chapter demonstrates the capability of two strains of Scenedesmus acutus to grow in ultra-high-density (>10 g L-1 dry weight biomass) cultures in flat panel photobioreactors for year-round production in the desert Southwest with record volumetric biomass productivity. The advantages and efficiency of high-density cultivation are discussed. The second chapter focuses on uptake and utilization of the volatile components of wastewater: ammonia and carbon dioxide. Scenedesmus acutus was cultured on wastewater from both municipal and agricultural origin and was shown to perform significantly better on flue gas as compared to commercial grade CO2 and just as well on waste nutrients as the commonly used BG-11 laboratory culture media, all while producing up to 50% lipids of the dry weight biomass suitable for use in biodiesel. The third chapter evaluates the feasibility of using gravity sedimentation for the harvesting of the difficult-to-separate Scenedesmus acutus green algae biomass followed by microfluidization to disrupt the cells. Lipid-extracted biomass was then studied as a fertilizer for plants and shown to have similar performance to a commercially available 4-6-6 fertilizer. Based on the work from these three chapters, a summary of modifications are suggested to help current and future microalgae companies be more competitive in the marketplace with traditional agricultural commodities. / Dissertation/Thesis / Doctoral Dissertation Molecular and Cellular Biology 2019

The atmospheric CO2 concentration is rising. These higher CO2 concentrations will certainly affect growth but may also affect nutrient responses of crop plants. The overall aim of this thesis was to investigate the effect of elevated CO2 on the nitrogen and phosphorus nutrition of two different crop species: wheat and cotton. Once establishing the new nutrient requirements of these crop species at elevated CO2 the study focussed on the interactive effects of elevated CO2 and N supply on growth and leaf N concentrations in cotton and wheat, carbohydrate metabolism, source:sink interactions and photosynthetic enzyme levels in the two species. Having established these effects, the role of N supply in controlling source:sink effects on Rubisco concentration were studied at the molecular level in wheat. Finally, some implications of the effect of elevated CO2 on wheat were examined, specifically on the quality of flour produced from wheat grain grown at elevated CO2. The foliar nitrogen concentrations, required to produce maximum shoot growth (critical concentrations) were lower at elevated CO2 while the critical phosphorus concentrations were higher. / Doctor of Philosophy (PhD)

Synechocystis PCC 6803 is a photosynthetic bacterium that has the potential to make bioproducts from carbon dioxide and light. Biochemical production from photosynthetic organisms is attractive because it replaces the typical bioprocessing steps of crop growth, milling, and fermentation, with a one-step photosynthetic process. However, low yields and slow growth rates limit the economic potential of such endeavors. Rational metabolic engineering methods are hindered by limited cellular knowledge and inadequate models of Synechocystis. Instead, inverse metabolic engineering, a scheme based on combinatorial gene searches which does not require detailed cellular models, but can exploit sequence data and existing molecular biological techniques, was used to find genes that (1) improve the production of the biopolymer poly-3-hydroxybutyrate (PHB) and (2) increase the growth rate. A fluorescence activated cell sorting assay was developed to screen for high PHB producing clones. Separately, serial sub-culturing was used to select clones that improve growth rate. Novel gene knock-outs were identified that increase PHB production and others that increase the specific growth rate. These improvements make this system more attractive for industrial use and demonstrate the power of inverse metabolic engineering to identify novel phenotype-associated genes in poorly understood systems. / Singapore-MIT Alliance (SMA)

Helicosporidia are single-celled obligate endoparasites of invertebrates. They have a unique morphology and infection strategy, which make them unlike any other eukaryote. Molecular data were produced to clarify their phylogenetic relationship and to examine the evolution of their cryptic plastid. Phylogenetic analyses of 69 ribosomal proteins identified from an expressed sequence tag (EST) library showed that Helicosporidia are derived green algae and more specifically, are related to the trebouxiophyte algae. An obligate parasitic lifestyle is rare among plant and algal groups, and because Helicosporidium possesses no pigments and no chloroplast-like structure has been identified, photosynthetic ability has presumably been lost in this organism. I sought to examine the role that a relict plastid might play in Helicosporidium. I identified ESTs of 20 putatively plastid-targeted enzymes that are involved in a wide variety of metabolic pathways. As expected, no components of photosynthesis were found, but components of other metabolic pathways including sulfur metabolism and fatty acid, isoprenoid and heme biosynthesis suggest that Helicosporidium retains its plastid for these functions. The complete plastid genome of this species of Helicosporidium was sequenced and revealed only four protein-coding genes not involved in transcription or translation, with two of these confirming the metabolic functions suggested by the nuclear-encoded, plastid-targeted genes identified from the ESTs. In addition, the Helicosporidium plastid genome is one of the smallest known (37.5 kb). Its reduced size results from loss of many genes commonly found in plastids of other plants and algae (including all proteins that function in photosynthesis), elimination of duplicated genes and redundant tRNA isoacceptors, and minimization of intergenic spaces. The Helicosporidium plastid genome is also highly structured, with each half of the circular genome containing nearly all genes on one strand. Both the structure and content of the plastid genome and the deduced function of the organelle show parallels with the relict plastid found in the malaria parasite, Plasmodium falciparum. These unrelated organisms each evolved from photosynthetic ancestors, and the convergence in form and function of their relict plastids suggest that common forces shape plastid evolution, following the switch from autotrophy to parasitism.

Helicosporidia are single-celled obligate endoparasites of invertebrates. They have a unique morphology and infection strategy, which make them unlike any other eukaryote. Molecular data were produced to clarify their phylogenetic relationship and to examine the evolution of their cryptic plastid. Phylogenetic analyses of 69 ribosomal proteins identified from an expressed sequence tag (EST) library showed that Helicosporidia are derived green algae and more specifically, are related to the trebouxiophyte algae. An obligate parasitic lifestyle is rare among plant and algal groups, and because Helicosporidium possesses no pigments and no chloroplast-like structure has been identified, photosynthetic ability has presumably been lost in this organism. I sought to examine the role that a relict plastid might play in Helicosporidium. I identified ESTs of 20 putatively plastid-targeted enzymes that are involved in a wide variety of metabolic pathways. As expected, no components of photosynthesis were found, but components of other metabolic pathways including sulfur metabolism and fatty acid, isoprenoid and heme biosynthesis suggest that Helicosporidium retains its plastid for these functions. The complete plastid genome of this species of Helicosporidium was sequenced and revealed only four protein-coding genes not involved in transcription or translation, with two of these confirming the metabolic functions suggested by the nuclear-encoded, plastid-targeted genes identified from the ESTs. In addition, the Helicosporidium plastid genome is one of the smallest known (37.5 kb). Its reduced size results from loss of many genes commonly found in plastids of other plants and algae (including all proteins that function in photosynthesis), elimination of duplicated genes and redundant tRNA isoacceptors, and minimization of intergenic spaces. The Helicosporidium plastid genome is also highly structured, with each half of the circular genome containing nearly all genes on one strand. Both the structure and content of the plastid genome and the deduced function of the organelle show parallels with the relict plastid found in the malaria parasite, Plasmodium falciparum. These unrelated organisms each evolved from photosynthetic ancestors, and the convergence in form and function of their relict plastids suggest that common forces shape plastid evolution, following the switch from autotrophy to parasitism.

Salt stress is one of the major problems in the present world’s agriculture. Plants encounter drought stress even in the availability of water because of osmotic imbalance in the cell due to excess salts. Plants avoid water uptake, which in turn decreases the photosynthetic activity. In this work, we measured the effect of salt stress on three accessions of Arabidopsis thaliana (Columbia (Col-0), Landsberg erecta (Ler-0), Wassilewskija (Ws-4)) by subjecting the plants to stress with 0-150 mM NaCl followed by recovery. The impact of the stress was clearly observed in all three accessions during stress and recovery period. Chlorophyll content in leaves decreased with increasing salt concentration. Proline levels increased during salt stress conditions. Non-photochemical quenching and PSII activity slightly decreased under stress conditions. Salt treated plants showed slow acidification of lumen with delayed Non-photochemical quenching in recovery phase. Ler-0 was the most sensitive ecotype to salt stress followed by Ws-4 and Col-0.