Cyanobacteria produce a variety of secondary metabolites which possess amongst others antifungal, antibacterial, and antiviral properties. Being primary producers they are also a vital component within the food web. However, certain strains also produce toxic metabolites such as the hepatotoxins microcystin (MC) and nodularin (NOD). Their toxicity in combination with the increasing global occurrence has resulted in a drinking water guideline limit of 1 μg L-1 being issued by the World Health Organisation (WHO). However, these toxins are not only present in water, but can be accumulated by fish and shellfish. Currently, no regulations regarding cyanotoxin contaminated seafood has been established despite similar toxicity to routinely monitored marine toxins such as domoic acid (DA). To facilitate regular monitoring, a high performance liquid chromatography photo diode array (HPLC-PDA) analysis method for the detection of DA was optimised to enable the simultaneous detection of DA and nine cyanotoxins. This method was then utilised to determine cyanotoxin concentration in laboratory cyanobacteria strains. To assess the accumulation and depuration of cyanotoxins in the common blue mussel Mytilus edulis, three feeding trials were performed. During these, mussels were exposed to two cyanobacteria strains, Nodularia spumigena KAC66, Microcystis aeruginosa PCC 7813, both individually and simultaneously. A rapid dose dependent accumulation of cyanotoxins was observed with maximum concentration of 3.4 -17 μg g-1 ww accumulated by M. edulis, which was followed by a much slower depuration observed. During the final feeding trial, with N. spumigena KAC 66 and M. aeruginosa PCC7813, cyanotoxins were still detectable following 27 days of depuration. Mortality in all studies was 7% or less indicating that most mussels were unaffected by the maximum dose of 480 μg L-1 NOD (feeding study 1), 390 μg L-1 MC (feeding study 2), or 130 μg L-1 total cyanotoxins (feeding trial 3), respectively. Mortality in negative control tanks was lower throughout all three feeding trials ( < 1 - 2.6%). Consumption of a typical portion size (20 mussels) would result in ingestion of cyanotoxins at levels significantly higher than the WHO recommended tolerable daily intake (TDI) of 2.4 μg NOD and/or MCs for a 60 kg adult. This value was exceeded not only during the exposure period (maximum levels 270 - 1370 μg cyanotoxins per 20 mussels), but also at the end of the depuration period 39-600 μg cyanotoxins per 20 mussels. These results illustrated that cyanotoxin monitoring of seafood should be considered not only during, but also following bloom events. In an attempt to investigate the cyanotoxin budget of the experimental system, not only mussels, but cyanobacteria cultures, the tank water, and the mussel faeces were also analysed for their cyanotoxin content. Results showed that large quantities of MCs and NOD were unaccounted for during all exposure trials. The combined effect of cyanotoxin metabolism in M. edulis, biotic and/or abiotic degradation, protein binding, and losses during the extraction and analysis were thought to have contributed to the unaccounted cyanotoxin fraction. Mussel flesh was analysed for the presence of glutathione or cysteine conjugates, however, there was no evidence of their occurrence in the samples tested. Due to these discrepancies in the toxin budget of the system, the introduction of correction factors for the analysis of cyanotoxins in M. edulis was suggested in order to protect the general public.

Submitted by Fabricia Fialho Reginato (fabriciar) on 2015-07-16T01:20:30Z No. of bitstreams: 1 JulianaElisa.pdf: 797306 bytes, checksum: 6777b1d4dc1bed955412dbeba92c4279 (MD5) / Made available in DSpace on 2015-07-16T01:20:30Z (GMT). No. of bitstreams: 1 JulianaElisa.pdf: 797306 bytes, checksum: 6777b1d4dc1bed955412dbeba92c4279 (MD5) Previous issue date: 2012 / CNPQ – Conselho Nacional de Desenvolvimento Científico e Tecnológico / Atualmente, há uma necessidade crescente pela busca por alternativas viáveis energeticamente, de baixo custo e sustentáveis que substituam o uso de combustíveis fósseis. Uma opção é a produção de biomassa energética através de microalgas, considerada uma alternativa limpa em relação a outras que demandam amplas áreas para cultivo e são geradoras de impactos ambientais. Nesse contexto, a pesquisa teve como objetivo avaliar a influência da temperatura e nutrientes no teor de lipídios de culturas de espécies de microalgas de água doce, visando o uso destes lipídios para a produção de biodiesel. Para isso, duas cepas de Monoraphidium contortum, uma cepa contendo as espécies de Chlorella vulgaris e Desmodesmus quadricauda e outra de Microcystis aeruginosa, foram mantidas em laboratório, por seis dias, em cinco meios de cultura ASM-1 modificados (controle com altas concentrações de fosfato e nitrato; sem fosfato; com concentração intermediária não limitante de fosfato; sem nitrato; e o último com concentração intermediária não limitante de nitrato). Posteriormente, foram submetidas às temperaturas de 13 ºC, 25 ºC (controle) e 37 ºC, durante oito dias (n=3). A extração dos lipídios foi realizada no 1°, 4° e 8° dias de experimento, seguindo metodologia comumente utilizada, através de solventes a frio, com a mistura de metanol e clorofórmio. Não foi observada variação lipídica significativa entre as cepas (p>0,05). Em média, as maiores produções lipídicas totais foram observadas quando as cepas foram mantidas em 13 ºC e no meio com concentração intermediária não limitante de nitrato. Foi encontrada variação significativa no percentual lipídico total em relação ao fator dia em que foi realizada a extração (p<0,05). O percentual lipídico variou em função da concentração de biomassa algal (p<0,05). A cepa de Microcystis aeruginosa obteve altos teores lipídicos totais, quando mantida em meio de cultura com alteração das concentrações de nitrato e fosfato, contrariando os resultados encontrados na literatura. As cepas testadas podem ser consideradas como potenciais produtoras de lipídios, desde que o meio de cultura, a temperatura e o dia de extração lipídica sejam corretamente estabelecidos. Este estudo mostrou que manipulações de fatores determinantes podem induzir maior concentração lipídica, otimizando a produção total com vistas à utilização desta matéria-prima para o biodiesel. / Currently, there is a growing need for the search of viable alternative energy, affordable and sustainable substitute for fossil fuels. One option is the production of energetic biomass by microalgae, which is considered a clean alternative compared to others that require large areas for cultivation and that generate environmental impacts. In this context, the research aimed to evaluate the influence of temperature and nutrients on lipid content of cultured species of freshwater microalgae, in order to use these lipids for biodiesel production. For this purpose, two strains of Monoraphidium contortum, a strain containing Chlorella vulgaris and Desmodesmus quadricauda and other of Microcystis aeruginosa, were maintained in the laboratory for six days, in five culture media, modified ASM-1 (control, with high concentrations of phosphate and nitrate; Pdeficient; non-limiting concentration of phosphate; N-deficient; non-limiting nitrate). Later, they were exposed to temperatures of 13 ° C, 25 ° C (control) and 37 ° C for eight days (n = 3). The lipid extraction was performed on the 1st, 4th and 8th day of the experiment following the method commonly used with cold solvent (a mixture of methanol and chloroform). There was no significant lipid variation among strains (p> 0.05). On average, the highest total lipid yields were observed when the strains were maintained at 13 ° C and in the medium with intermediate concentration and nonlimiting nitrate. We found significant variation in total lipid percentage in relation to day that the extraction was performed (p<0,05). The lipid percentage varied depending on the concentration of algal biomass (p<0,05). The strain of Microcystis aeruginosa showed high total lipid content, when maintained in culture medium with abnormal concentrations of nitrate and phosphate, contrary to the literature data. The strains can be considered as potential lipid producers since the culture medium, the temperature and the time for extraction of lipids are correctly established. This study showed that manipulations of determining factors might induce higher lipid concentration, optimizing the total production in order to use this raw material for biodiesel.

Made available in DSpace on 2015-09-25T12:22:40Z (GMT). No. of bitstreams: 1 PDF - Erivanna Karlene dos Santos Oliveira.pdf: 2316361 bytes, checksum: a621cb80b0ca857fdd90fc5f90fbf6bf (MD5) Previous issue date: 2014-04-11 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / The presence of cyanobacteria in reservoirs, lakes and rivers is a global environmental health problem as a result ofanthropogenic eutrophication. These microorganisms hinder water purifiers, to produce taste and odor difficult to remove and the cells at high density clog the sand filters, reducing its time of use. In the conventional method of water treatment, coagulation, flocculation and sedimentation are essential in the removal of cells and determine the efficiency of the following operations. In this study we analyzed the efficiency of removal of intact cells of Microcystis aeruginosaand microcystin-LR in the stages of coagulation, flocculation and sedimentation, followed by filtration through sand filters with the use of coagulants ferric chloride and aluminum sulfate, evaluating four filtration rates: 15, 30, 50 and 100 m 3 .m -2 .d -1 at pH 7.5 and a dosage of 40 mg.L -1 tests with a bench-scale (Jartest) using treated water dechlorinated and adadded with a pureculture of M. aeruginosa with approximate final densityof 10 cel.mL -1 simulating a bloom. The parametres control were turbidity, apparent color and cell concentration. It was observed that both the efficiency rates at 15 and 30 m 1 had similar removal of apparent color, turbidity and cellconcentration (79%, 80% and 92% and 80%, 80,5% and 89,10% respectively for the coagulant ferric chloride. And for aluminum sulphate was the same for both rates removals of 81,48% for apparent color, turbidity to 83,25% and to 90,50% for cell concentration, since the rate for 30 m 1 removals were 83,33%, 83,25% and 89,71%, respectively. Nocell lysiswas observedandconsequently, no further increases in concentrationsofmicrocystin-LR cyanotoxinduring treatment was observed. / A presença de cianobactérias em reservatórios, lagos e rios é um problema mundial de saúde ambiental, em consequência da eutrofização antropogênica. Esses microrganismos dificultam a potabilização da água, por produzir gosto e odor difíceis de remover e suas células em alta densidade colmatam os filtros, diminuindo sua vida útil. No método convencional de tratamento de água, coagulação, floculação e sedimentação são fundamentais na remoção de células e determinam a eficiência das operações seguintes. Neste trabalho analisou-se a eficiência na remoção de células intactas de Microcystis aeruginosa e da microcistina-LR nas etapas de coagulação, floculação e sedimentação, seguido por filtração em filtro de areia com uso dos coagulantes cloreto férrico e sulfato de alumínio, avaliando quatro taxas de filtração de 15, 30, 50 e 100 m 3 .m , em pH 7,5 e dosagem de 40 mg.L -1 com testes em escala de bancada (Jartest) utilizando-se água tratada de torneira desclorada e adicionada de uma cultura pura de M. aeruginosa com densidade final aproximada de 10 5 cel.mL -1 simulando uma floração. Os parâmetros de referência foram turbidez, cor aparente e concentração celular. Observou-se que a eficiência, tanto com as taxas de 15 e 30 m 3 .m -2 .d -1 tiveram remoções semelhantes para as variáveis cor aparente, turbidez e concentração celular com 79%, 80% e 92%; e 80%, 80,5% e 89,10%, respectivamente para o coagulante cloreto férrico. Para o sulfato de alumínio e para ambas as taxas, houve a mesma reposta com remoções de 81,48% para cor aparente, 83,25% para turbidez e 90,50% para concentração celular, já para a taxa de 30 m3 .m -2 .d -1 as remoções foram de 83,33% , 83,25% e 89,71% respectivamente. Não foi observada lise celular e consequentemente, não houve aumentos, ao longo do tratamento, das concentrações da cianotoxina microcistina -LR.

The Laurentian Great Lake, Lake Erie is an invaluable global resource and its watershed is home to over 11 million people. The pressures placed on the lake because of this high population caused Lake Erie to experience numerous environmental problems, including seasonal hypoxia and harmful algal blooms. While these topics have been widely studied in Lake Erie for over 40 years a more nuanced understanding of the interaction between phytoplankton and nutrient is needed to properly address the problems continuing to face the lake. In this study we combine classical limnological and cell growth experiments with modern molecular biological techniques and microscopy to more completely describe the aquatic microbial ecology of the lake. We used an oxalate rinse technique to examine the surface absorbed P pool of the toxic cyanobacterium Microcystis aeruginosa grown under a range of P conditions, as well as the general Lake Erie plankton assemblage. Our results suggest that while Microcystis is plastic in its cellular P needs, the ratio of intracellular to extracellular P remains stable across growth conditions. We describe the effect of the phosphonate herbicide glyphosate on the Lake Erie phytoplankton community using laboratory cell growth studies, field microcosm experiments and PCR amplification of a gene implicated in the breakdown of this compound from the environment. Results from these experiments suggest that the presence of glyphosate can affect community structure in multiple ways and may explain areas of unexplained phytoplankton diversity in coastal areas of Lake Erie. We also show heterotrophic bacteria are likely critical to the breakdown of glyphosate and further illustrate that understanding the context of the larger microbial community is critical to understanding the ecology of the constituent members of the community. Finally, we investigate the activity of the phytoplankton community in winter months with a focus on diatoms abundant in Lake Erie under the ice. We show these diatoms are active and that the winter bloom is a likely source of carbon important to seasonal hypoxia formation. Together, these studies significantly enrich our understanding of how phytoplankton influence important ecological processes in Lake Erie.

Certain types of cyanobacteria have the potential to produce toxins including microcystin, a hepatotoxin. Toxic cyanobacterial blooms are becoming increasingly common worldwide. They are a concern in the Great Lakes and surrounding waters. In this study, Lake Ontario’s Bay of Quinte, Lake Erie’s Maumee Bay, and three reservoirs along the Grand River were studied. Environmental variables, cyanobacterial biomass inferred from the Fluoroprobe, and microcystin concentrations were measured. In 2005 the three reservoirs, Belwood Lake, Conestogo Lake, and Guelph Lake were sampled every two weeks from July to September. Belwood Lake was also sampled in October when a cyanobacterial bloom occurred. In 2006 the Bay of Quinte was sampled twice, in July and September, and Maumee Bay was sampled twice, in June and August. Physical variables measured included water transparency and temperature. All species of nitrogen (N) and phosphorus (P) were measured, along with extracted chlorophyll a and particulate carbon (C), N, and P. The distribution of chlorophyll and major algal groups throughout the water column was profiled in situ using a spectral fluorometer (Fluoroprobe).Variable fluorescence of phytoplankton was assessed using Pulse Amplitude Modulated (PAM) fluorometry to measure photosynthetic parameters. Phytoplankton counts were performed on selected samples from the Bay of Quinte and Maumee Bay. Total and dissolved microcystin were measured using the protein phosphatase inhibition assay (PPIA). PPIA was chosen over alternative detection methods because it is a functional assay that measures the level of microcystin in a sample via the amount of protein phosphatase inhibition that it exerts. This yields ecologically relevant data as protein phosphatase inhibition is the main mode of microcystin toxicity. The PPIA formulation used in our lab was based on variations in the literature that use unconcentrated water samples directly in the assay. The assay was optimized to employ both a higher and lower standard curve through the use of two enzyme concentrations. The lower enzyme concentration allowed the method detection limit to be decreased to 0.05 µg/L to accommodate our low-microcystin samples. In the Bay of Quinte, microcystin levels were higher in July 2006 (total mean=2.25 μg/L ) than in September 2006 (total mean=0.58 μg/L). In July a cyanobacterial bloom consisting of 97% Microcystis spp. was present. In September 83% of the cyanobacterial biomass was composed of Anabaena spiroides and only 8% was Microcystis spp. In the Bay of Quinte elevated microcystin concentrations were associated with higher soluble reactive P levels, lower seston C:P molar ratios, and lower total N. In Maumee Bay microcystin levels were higher in August 2006 (total mean= 4.45 μg/L) than they were in June 2006 (<0.05 μg/L). In August a cyanobacterial bloom consisting of 22% Microcystis spp. and 48% Aphanizomenon flos-aquae was observed. Higher microcystin concentrations in Maumee Bay were associated with decreased total N: total P molar ratios, increased total P, and decreased water transparency as measured by Secchi depth. Belwood Lake had the highest microcystin levels of the three reservoirs but only once exceeded the recommended World Health Organization concentration of 1.0 μg/L. Belwood Lake’s largest cyanobacterial bloom in October 2005 was accompanied by relatively low microcystin levels (<0.2 μg/L). Conestogo and Guelph lakes always had microcystin levels below 0.2 μg/L and 0.6 μg/L, respectively. In the Grand River reservoirs, increased microcystin concentrations were associated with higher chlorophyll a, higher light attenuation coefficients, lower total N, lower total N: total P molar ratios, higher C:P molar ratios, lower nitrate, higher cyanobacterial biomass, and higher total P. When data from the Bay of Quinte, Maumee Bay, and Grand River reservoirs were pooled, total microcystin had the most significant positive correlation with total P. Total microcystin and water temperature also had a significant positive correlation.

Certain types of cyanobacteria have the potential to produce toxins including microcystin, a hepatotoxin. Toxic cyanobacterial blooms are becoming increasingly common worldwide. They are a concern in the Great Lakes and surrounding waters. In this study, Lake Ontario’s Bay of Quinte, Lake Erie’s Maumee Bay, and three reservoirs along the Grand River were studied. Environmental variables, cyanobacterial biomass inferred from the Fluoroprobe, and microcystin concentrations were measured. In 2005 the three reservoirs, Belwood Lake, Conestogo Lake, and Guelph Lake were sampled every two weeks from July to September. Belwood Lake was also sampled in October when a cyanobacterial bloom occurred. In 2006 the Bay of Quinte was sampled twice, in July and September, and Maumee Bay was sampled twice, in June and August. Physical variables measured included water transparency and temperature. All species of nitrogen (N) and phosphorus (P) were measured, along with extracted chlorophyll a and particulate carbon (C), N, and P. The distribution of chlorophyll and major algal groups throughout the water column was profiled in situ using a spectral fluorometer (Fluoroprobe).Variable fluorescence of phytoplankton was assessed using Pulse Amplitude Modulated (PAM) fluorometry to measure photosynthetic parameters. Phytoplankton counts were performed on selected samples from the Bay of Quinte and Maumee Bay. Total and dissolved microcystin were measured using the protein phosphatase inhibition assay (PPIA). PPIA was chosen over alternative detection methods because it is a functional assay that measures the level of microcystin in a sample via the amount of protein phosphatase inhibition that it exerts. This yields ecologically relevant data as protein phosphatase inhibition is the main mode of microcystin toxicity. The PPIA formulation used in our lab was based on variations in the literature that use unconcentrated water samples directly in the assay. The assay was optimized to employ both a higher and lower standard curve through the use of two enzyme concentrations. The lower enzyme concentration allowed the method detection limit to be decreased to 0.05 µg/L to accommodate our low-microcystin samples. In the Bay of Quinte, microcystin levels were higher in July 2006 (total mean=2.25 μg/L ) than in September 2006 (total mean=0.58 μg/L). In July a cyanobacterial bloom consisting of 97% Microcystis spp. was present. In September 83% of the cyanobacterial biomass was composed of Anabaena spiroides and only 8% was Microcystis spp. In the Bay of Quinte elevated microcystin concentrations were associated with higher soluble reactive P levels, lower seston C:P molar ratios, and lower total N. In Maumee Bay microcystin levels were higher in August 2006 (total mean= 4.45 μg/L) than they were in June 2006 (<0.05 μg/L). In August a cyanobacterial bloom consisting of 22% Microcystis spp. and 48% Aphanizomenon flos-aquae was observed. Higher microcystin concentrations in Maumee Bay were associated with decreased total N: total P molar ratios, increased total P, and decreased water transparency as measured by Secchi depth. Belwood Lake had the highest microcystin levels of the three reservoirs but only once exceeded the recommended World Health Organization concentration of 1.0 μg/L. Belwood Lake’s largest cyanobacterial bloom in October 2005 was accompanied by relatively low microcystin levels (<0.2 μg/L). Conestogo and Guelph lakes always had microcystin levels below 0.2 μg/L and 0.6 μg/L, respectively. In the Grand River reservoirs, increased microcystin concentrations were associated with higher chlorophyll a, higher light attenuation coefficients, lower total N, lower total N: total P molar ratios, higher C:P molar ratios, lower nitrate, higher cyanobacterial biomass, and higher total P. When data from the Bay of Quinte, Maumee Bay, and Grand River reservoirs were pooled, total microcystin had the most significant positive correlation with total P. Total microcystin and water temperature also had a significant positive correlation.

Light is the primary source of energy in most of earth's ecosystems . In freshwater ecosystems the major interacting factors that determine the abundance and species composition of planktonic phototrophs, the primary utilizers of light, are nutrients, temperature and light. With increasing eutrophication and declining geographical latitude, nutrient availability becomes in excess of the organisms' requirements, water temperature is more favourable for growth, and community structure depends to a greater extent on light availability. This study focuses on the population dynamics of the bloom-forming cyanobacterium Microcystis aeruginosa Kutz. emend. Elenkin in subtropical Hartbeespoort Dam, South Africa. The objectives of the study were: to investigate the annual cycle, and the factors leading to the dominance and success of the cyanobacterium in this hypertrophic, warm monomictic lake, where light availability is the major factor limiting phytoplankton growth rates; to study the surface blooms and ultimately hyperscums that this species forms; and to assess the ecological significance of hyperscums. A 4. 5-years field study of phytoplankton abundance and species composition in relation to changes in the physical environment, was undertaken. The hypothesis was that M. aeruginosa dominated the phytoplankton population (> 80 % by volume) up to 10 months of every year because it maintained itself within shallow diurnal mixed layers and was thus ensured access to light. It was shown that wind speeds over Hartbeespoort Dam were strong enough to mix the epilimnion (7 - 18 m depth) through Langmuir circulations only 12 % of the time. At other times solar heating led to the formation of shallow ( < 2 m) diurnal mixed layers (Z[1]) that were usually shallower than the euphotic zone (Zeu; x = 3.5 m), while the seasonal mixed layer (zrn) was always deeper than Zeu. From the correspondence between vertical gradients of chlorophyll a concentrations and density gradients, when M. aeruginosa was dominant, it was implied that this species maintained the bulk of its population within Z[1]. Under the same mixing conditions non-buoyant species sank into dark layers. These data point out the importance of distinguishing between Zrn and Z[1], and show the profound effect that the daily pattern of Z[1], as opposed to the seasonal pattern of Zrn can have on phytoplankton species composition Adaptation to strong light intensities at the surface was implicated from low cellular chlorophyll a content (0.132 μg per 10[6] cells) and high I[k ](up to 1230 μE m⁻² S¯¹). Ensured access to light, the postmaximum summer populations persisted throughout autumn and winter, despite suboptimal winter temperatures, by sustaining low losses. Sedimentation caused a sharp decline of the population at the end of winter each year and a short ( 2-3 months) successional episode follCMed, rut by late spring M. aeruginosa. was again dominant. The mixing regime in Hartbeespoort Dam and the buoyancy mechanism of M. aeruginosa led to frequent formation of surface bloons and ultimately hyperscums. Hyperscums were defined as thick (decimeters), crusted, buoyant cyanobacterial mats, in which the organisms are so densely packed that free water is not evident. In Hartbeespoort Dam in winter M. aeruginosa formed hyperscums that measured up to 0.75 m in thickness, covered more than a hectare, contained up to 2 tonnes of chlorophyll a, and persisted for 2 - 3 monnths. Hyperscum formation was shown to depend upon the coincidence of the following preconditions: a large, pre-existing standing crop of positively buoyant cyanobacteria; turbulent mixing that is too weak to overcome the tendency of the cells to float, over prolonged periods (weeks); lake morphometry with wind-protected sites on lee shores; and high incident solar radiation. The infrequent occurrence of hyperscums can be attributed to the rare co-occurrence of these conditions. Colonies in the hyperscum were arranged in a steep vertical gradient, where colony compaction increased exponentially with decreasing distance form the surface. This structure was caused by evaporative dehydration at the surface, and by the buoyancy regulation mechanism of M. aeruginosa., which results with cells being unable to lose boyancy when deprived access to light from above. The mean chlorophyll a concentration and water content were 3.0 g 1¯¹ and 14 % at the surface crust, 1.0 g 1¯¹ and 77 % at a few mm depth, and 0.3 g 1¯¹ and 94 % at 10 cm depth, where M. aeruginosa cell concentration exceeded 109 ml¯¹. A consequence of the high cell and pigment concentrations was that light penetrated only 3 mm or less, below which anaerobic, highly reduced conditions developed. Nutrient concentrations in hyperscum interstitial water, collected by dialysis, increased dramatically with time (phosphate: 30-fold over 3 months; ammonia: 260-fold). Volatile fatty acids, intermediate metabolites in anaerobic decomposition processes, were present. Gas bubbles trapped within the hyperscum contained methane (28 %) , and CO[2] (19 %), the major end products of anaerobic decomposition, and no oxygen. The structure and function of M. aeruginosa in hyperscum was examined in relation to the vertical position of colonies and the duration of exposure to hyperscum condition. Colonies and cells collected from 10 em depth in the hyperscum were similar in their morphology (light and fluorescent microscopy) and ultrastructure (transmission and scanning electron microscopy) to those of colonies from surface blooms in the main basin of the lake. With declining depth over the uppermost 10 mm of the hyperscum cells appeared increasingly dehydrated, decomposed and' colonized by bacteria. studies employing microelectrode techniques demonstrated that photosynthetic activity of colonies at the surface of a newly accumulated hyperscum rapidly photoinhibited, substrate-limited, and then ceased within hours of exposure to light intensities > 625 μE m⁻² S¯¹. Photooxidative death followed. The dead cells dehydrated to form the dry crust, from underneath. and space was thus created for colonies rising Cells collected from 10 cm depth retained their photosynthetic capacity ([14]C-uptake experiments) throughout the hyperscum season, although a considerable decline in this capacity was noted over the last (third) month. Altogether the data indicated that spatial separation developed within the hyperscum, between a zone at the surface of lethal physical conditions, a zone beneath the surface of stressful and probably lethal chemical conditions, and a deeper zone of more moderate conditions, which nevertheless, deteriorated after 2 - 3 months. A conceptual model describing the fate of a colony entering a hyperscum was then proposed. According to this model, a colony that arrives below a hyperscum and is not carried away by currents, becomes over-buoyant in the dark and floats into the bottom of the hyperscum. With time it migrates towards, due to its own positive buoyancy, the buoyancy of colonies rising from underneath, and the collapse of cells at the top. It survives in the dark, anaerobic environment by maintaining low levels of basal metabolism while utilizing stored reserves. Depending on weather conditions, the colony mayor may not remain within the hyperscum long enough to reach the zone of decomposition near the surface, where it would die. With the aging of the hyperscum and the accumulation of trapped decomposition products, the zone of decomposition expands. Thus, a hyperscum is essentially a site of a continuous cycle of death and dehydration at the surface and upward migration of colonies from below to replace those that died, although not all colonies entering the hyperscum necessarily reach the lethal zone. The formation of hyperscums was shown to have no major influence on the annual cycle of M. aeruginosa in Hartbeespoort Dam. The seasonality of increase and decline of the planktonic population was similar from year to year, irrespective of whether or not hyperscums formed. The phenomenon of hyperscums demonnstrated that, as Reynolds and Walsby (1975) claimed, thick cyanobacterial water-blooms do form incidentally and have no vital function in the biology of the organism. water temperature did have a major effect on the annual cycle of this species in Hartbeespoort Dam. In temperate lakes the low water temperatures in autumn and winter (<10° C) cause M. aeruginosa to lose its ability to regain buoyancy in the dark, and consequently it sinks to bottom sediments. The higher ( > l2°C) minimum winter temperature in Hartbeespoort Dam leads to the maintenance of a relatively large residual planktonic population throughout the winter. Unlike the case in temperate lakes, the long-term survival of M. aeruginosa in warm-water lakes probably does not depend on winter benthic stocks for the provision of an inoculum for the following growth season. / Thesis (Ph.D.)-University of Natal, Pietermaritzburg, 1987.

Made available in DSpace on 2014-06-11T19:23:02Z (GMT). No. of bitstreams: 0 Previous issue date: 2011-02-17Bitstream added on 2014-06-13T19:28:52Z : No. of bitstreams: 1 santos_vp_me_rcla.pdf: 692384 bytes, checksum: 87577700198a680d7662c5fe0990efe7 (MD5) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / As cianobactérias apresentam uma ampla variabilidade fenotípica e ecológica. Porém, esta variabilidade, muitas vezes, não corresponde à sua diversidade genética. Assim, o uso de marcadores moleculares é fundamental para os estudos de filogenia neste grupo. Entretanto, a filogenia molecular enfrenta um desafio na seleção dos marcadores devido à ocorrência relativamente frequente da transferência de genes de forma lateral entre os procariotos. Em cianobactérias os marcadores dos espaçadores dos genes ribossomais (16S-23S-ITS rDNA) e do operon da ficocianina (PC-IGS) estão entre os mais utilizados nestes estudos. Contudo, alguns trabalhos sugerem que o PC-IGS possa ter sito transferido lateralmente em sua história evolutiva. A identificação de morfoespécies dos gêneros Microcystis e Geitlerinema é baseada em caracteres morfológicos que em geral não correspondem à sua variabilidade genética. Com o objetivo de investigar a transferência lateral do operon da ficocianina em Geitlerinema e Microcystis, foram obtidas e comparadas árvores filogenéticas de ambas espécies baseadas nos marcadores PC-IGS e 16S-23S-ITS rDNA. As topologias das árvores obtidas para ambos os marcadores foram muito semelhantes e indicaram que o PC-IGS é estável e indicado para os estudos de taxonomia e filogenia de linhagens de Geitlerinema e Microcystis. Assim, hipótese inicial de transferência lateral foi refutada. Algumas linhagens tiveram seu posicionamento divergente entre um marcador e outro, o que ressalta a importância do uso de mais de um marcador em estudos de filogenia. O marcador PC-IGS apresentou melhor desempenho que 16S-23S-ITS rDNA. As árvores filogenéticas de Geitlerinema baseadas em ambos os marcadores indicaram a ocorrência de espécies crípticas dentre as linhagens estudadas e corroboraram que G. amphibium e G. unigranulatum devem ser consideradas sinonímias... / Cyanobacteria show a wide phenotypic and ecological variability, but frequently this variability does not correspond to their genetic variation. Therefore, the use of molecular markes is critical for phylogenetic studies in this group. At the same time, the selection of molecular markers represents a challenge for the molecular phylogeny due to the horizontal gene transfer, witch is a relatively common process among the prokaryotes. In cyanobacteria, makers for the ribosomal genes spacer (16S-23S-ITS rDNA) and for the phycocyanin operon spacer (PC-IGS) are among of the most used for phylogeny. However, some studies suggest that the PC-IGS marker may have been horizontally transferred during its evolutionary history. The identification of the morphospecies from the genus Microcystis and Geitlerinema is based in their morphological characters, but they generally do not correspond to genetic variability. In order to investigate the possibility of horizontal transfer of the phycocyanin operon in Microcystis and Geitlerinema, phylogenetic trees based on the PC-IGS and 16S- 23S-ITS rDNA were generated and compared. The topologies obtained for both markers were very similar, indicating that the PC-IGS marker is stable and suitable for taxonomical and phylogenetic studies in Microcystis and Geitlerinema. Therefore, the initial hypothesis of horizontal transfer was rejected. Some strains were found to have divergent positions between the trees based on the two molecular markes, witch highlights the importance of using more than one marker in phylogenetic studies. The PC-IGS marker performed better than 16S-23SITS rDNA. The Geitlerinema phylogenetic trees based on both markers indicated the occurrence of cryptic species among the strains and corroborated that G. amphibium and G. unigranulatum should be treated as synonyms. The phylogenetic tree based on PC-IGS formed a monophyletic clade... (Complete abstract click electronic access below)

Made available in DSpace on 2016-06-02T19:31:47Z (GMT). No. of bitstreams: 1 2600.pdf: 865519 bytes, checksum: 4d45e3ac1d33d455fb108ecff59f5481 (MD5) Previous issue date: 2009-07-31 / Solar radiation can change DOM in aquatic environments making compounds more labile or recalcitrant to biodegradation. Thus, solar radiation, especially UV, indirectly interfere in the microbial community in order to increase or decrease their growth. The objective was to verify the interference of UV-A + B on the bioavailability of DOM separated into different fractions (molecular weight), from release of cells in stationary phase of M. aeruginosa. DOM released from cyanobacteria was separated by tangential ultra-filtration and these were inoculated bacterial community of Barra Bonita reservoir. Bacterial community dynamic was monitored by measures of density and biomass and bacterial consumption of carbon from DOM by measures of DOC. Frequency of bacterial morphotypes were also calculated. All three fractions of DOM investigated were highly available to the bacterial community. The fraction of cultures with DOM<3 kDa and DOM>30 kDa demonstrated to be more vulnerable to UV radiation, corroborated by many statistical differences found for bacterial density during 27 days of incubation. At 15 days of incubation, the cultures with irradiated DOM>30 kDa showed a second peak in bacterial density and biomass that can be understood as an event of succession in bacterial community. Changes in relative frequency of bacterial morphotypes over incubation period suggest an episode of succession during the different stages of degradation of DOM. It was suggested that the majority of DOC released from a bloom of cyanobacteria is rapidly mineralized. Furthermore, the constant blooms, or ever high biomass of M. aeruginosa during the year, mean that such bacterial consortia are always available. / A radiação solar pode alterar a MOD em um ambiente aquático tornando os compostos mais lábeis ou recalcitrantes à biodegradação. Desta forma, a radiação solar, principalmente a radiação UV, interfere indiretamente na comunidade microbiana de forma a incrementar ou diminuir seu crescimento. O objetivo deste trabalho foi verificar a interferência da radiação UV-A+B na biodisponibilidade da MOD separada em diferentes frações (massa molecular), oriundas da liberação de células na fase estacionária de Microcystis aeruginosa. A MOD liberada pela cianobactéria foi separada por ultra-filtração tangencial e a estas foram inoculadas a comunidade bacteriana do reservatório de Barra Bonita. A dinâmica da comunidade bacteriana foi acompanhada por medidas de densidade e biomassa bacteriana e o consumo de carbono das frações de MOD por medidas de COD. Também foram calculadas as frequências dos morfotipos bacterianos. Todas as três frações de MOD investigadas apresentaram-se altamente disponíveis à comunidade bacteriana. As culturas com fração de MOD<3 kDa e MOD>30 kDa revelaram-se mais vulneráveis à radiação UV, fato corroborado pelas numerosas diferenças estatísticas encontradas para as densidades bacterianas durante os 27 dias de incubação. Aos 15 dias de incubação, as culturas com fração de MOD>30 kDa irradiada apresentaram um segundo pico na densidade e biomassa bacteriana que pode ser interpretado como um evento de sucessão na comunidade bacteriana. As mudanças na frequência relativa dos morfotipos bacterianos ao longo do período de incubação sugerem um episódio de sucessão durante as distintas fases de degradação da MOD. Sugeriu-se que a maior parte do COD liberado por um bloom desta cianobactéria é mineralizado rapidamente. Além disso, com os constantes blooms, ou a sempre alta biomassa de M. aeruginosa ao longo do ano, inferem que tais consórcios bacterianos estejam sempre disponíveis.

Cyanobacteria (“blue-green algae”) are known to produce a diverse repertoire of biologically active secondary metabolites. When associated with so-called “harmful algal blooms”, particularly in freshwater systems, a number of these metabolites have been associated - as “toxins”, or commonly “cyanotoxins” - with human and animal health concerns. In addition to the known water-soluble toxins from these genera (i.e. microcystins, cylindrospermopsin, and saxitoxins), our studies have shown that there are metabolites within the lipophilic extracts of these strains that inhibit vertebrate development in zebrafish embryos. Following these studies, the zebrafish embryo model was implemented in the bioassay-guided purification of four isolates of cyanobacterial harmful algal blooms, namely Aphanizomenon, two isolates of Cylindrospermopsis, and Microcystis, in order to identify and chemically characterize the bioactive lipophilic metabolites in these isolates. We have recently isolated a group of polymethoxy-1-alkenes (PMAs), as potential toxins, based on the bioactivity observed in the zebrafish embryos. Although PMAs have been previously isolated from diverse cyanobacteria, they have not previously been associated with relevant toxicity. These compounds seem to be widespread across the different genera of cyanobacteria, and, according to our studies, suggested to be derived from the polyketide biosynthetic pathway which is a common synthetic route for cyanobacterial and other algal toxins. Thus, it can be argued that these metabolites are perhaps important contributors to the toxicity of cyanobacterial blooms. In addition to the PMAs, a set of bioactive glycosidic carotenoids were also isolated because of their inhibition of zebrafish embryonic development. These pigmented organic molecules are found in many photosynthetic organisms, including cyanobacteria, and they have been largely associated with the prevention of photooxidative damage. This is the first indication of these compounds as toxic metabolites and the hypothesized mode of action is via their biotransformation to retinoids, some of which are known to be teratogenic. Additional fractions within all four isolates have been shown to contain other uncharacterized lipophilic toxic metabolites. This apparent repertoire of lipophilic compounds may contribute to the toxicity of these cyanobacterial harmful algal blooms, which were previously attributed primarily to the presence of the known water-soluble toxins.