Thesis (M.S.)--University of Wisconsin--Madison. / Typescript. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 107-119).

Neither the ecological role nor the metabolic function of microcystin is known. Cellular microcystin concentrations correlate to cellular nitrogen status for a given environmental phosphorous concentration and specific growth rate. Microcystin production is enhanced when the rate of nitrogen accumulation exceeds the relative specific growth rate and/or when cellular N:C ratios exceed the Redfield ratio as a function of reduced carbon fixation, suggesting enhanced production of microcystin under carbon stress. Additionally, a strong correlation between medium phosphate and carbon fixation, and the negative correlation between medium phosphate and microcystin combined with the cellular localization of microcystin in thylakoids supports a possible role for microcystin in enhancement of photosynthesis. Batch cultures of both Microcystis aeruginosa PCC7806 and a mcyA- knockout mutant of PCC7806 were therefore cultured at different light intensities and media treatments, so as to vary cellular N:C ratios and concentrations, and sampled for analysis of microcystin concentration, cell numbers and residual medium nitrates. Inter-strain differences in photosynthetic electron transfer rates and levels were monitored using a Hansatech PEA fluorometer and compared to cellular microcystin concentrations. An enhanced survival was observed at high light, where the toxic strain survived while the nontoxic strain became chlorotic. A strong correlation (r2 = 0.907, p< 0.001, N=22) between microcystin concentration and growth rate was observed at high light conditions. No such advantage was observed at optimal or low-light conditions and media composition had no significant effect on the relationship between toxicity and survival at high light. PCC7806 showed elevated PI(abs) values compared to the mcyA knockout strain, which indicates an increased stability of PSII. A strong correlation between PI(abs) and microcystin (r = 0.88, p< 0.005, N=15) was observed for cultures grown in modified BG11 containing 25 mM under continuous illumination of 37 μmol of photons m-2.s-1. No correlation was observed between PI(abs) and microcystin for the other treatments. The toxin producer had significantly higher values for density of active reaction centers and ii quantum efficiency compared to the mutant. A decrease in F0 in the mutant suggests degradation of the phycobiliproteins, whereas PCC7806 didn’t show a significant decrease in F0 Data indicate that microcystins play a role in photosynthesis by preventing chlorosis in saturating light conditions either by enhancing the redox stability of the phycobiliproteins or PS II, thus preventing photooxidation.

No description available.

Thesis (Ph. D.)--University of Wisconsin--Madison, 1959. / Typescript. Abstracted in Dissertation abstracts, v. 20 (1959) no. 2, p. 470. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 90-92).

This study reports the findings on the effect of Microcystin-LR (MCLR) on the gastrointestinal tract cells of mice and on two different cell lines, Caco2 and MCF-7. The cyanobacterium Microcystis aeruginosa produces the potent toxin, MCLR. This toxin has been implicated in a number of cases of ill-health. It was decided to investigate whether microcystin-LR induced apoptosis in the gastrointestinal tract of mice and also which possible mechanisms were involved in the induction in vitro. Balb/c mice were given a 75% LD50 intraperitoneal dose of pure microcystin -LR and sacrificed at 8, 16, 24 and 32 hours post-exposure. The small intestinal sections were stained with haematoxylin and eosin and examined for apoptotic cells. There was a time-dependent increase in the number of apoptotic cells with most in the duodenum and the jejunum. No change in glycogen content was evident at 24 hours post exposure when PAS-stained sections were examined. To determine that microcystin was the agent responsible for the changes, fluoroscein isothiocyanate (FITC) immunostaining for the toxin was done on the sections. Apoptosis in vitro was investigated in Caco2, a cell line that behaves like normal enterocytes when the cells are differentiated at confluency, and MCF-7, a breast cancer cell line deficient in pro-caspase-3, cells by 3-[dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) and lactate dehydrogenase (LDH) assays and by staining with DAPI and Rhodamine 123. MCLR exposure induced apoptosis, as seen in decreased cell viability and increased leakage of LDH, as well as mitochondrial damage shown by Rhodamine staining. The MCF-7 cells, deficient in pro-caspase-3, and Caco2 cells did not show cleavage of poly(ADP)ribose polymerase (PARP) after exposure to 50μM MCLR after 72 hours exposure. Both micro- and milli-calpain activity was however significantly increased in both cell lines exposed to the toxin. There was a significant increase in H2O2, one of the key reactive oxygen species, production during the first 30 minutes that the cells were exposed to 50 mM MCLR.

A module and an inoculation technique were developed that would allow for the efficient immobilization of Microcystis aeruginosa PCC7806 on a synthetic membrane. A variety of module types, membranes (ceramic, tubular polyethersulfone and externally skinless polyethersulfone capillary membrane), and methods of immobilization (adsorption, pressure filtration and a developed technique that involved drying a cell slurry on a membrane) were assessed. The morphological properties that affected the immobilization of Microcystis aeruginosa PCC7806, as well as the effects of immobilization upon cell morphology were assessed. Cells in the stationary growth phase, which had a well-developed extra-cellular polysaccharide layer and no gas vesicles, were optimal for immobilization. Microcystin production under immobilized conditions was assessed under different nitrate concentrations, light intensities, biofilm thickness and immobilization times. Additional work included assaying for Microcystin production of two airlift-grown cultures under a high light intensity and complete nutrient deprivation and the inoculation of a ceramic membrane. An immunological technique was used to elucidate where toxin production was greatest within a biofilm immobilized upon an externally skinless polyethersulfone capillary membrane. The externally skinless polyethersulfone capillary membrane was evaluated to assess homogeneity and the physical differences between membrane batches that led to the erratic, incomplete biofilm formation, as a biofilm of a constant thickness could not be immobilized. Microcystis aeruginosa PCC7806 was exposed to a variety of solvents in order to permeabilize the cyanobacteria, as that would have enabled a truly continuous extraction process for the metabolite. FDA hydrolysis had to be optimized in order to use it as an indicator of cell viability. In addition a single-step extraction of Microcystin was attempted using live bacteria. A capillary membrane module, containing the externally skinless polyethersulfone capillary membrane, inoculated using pressure filtration, was the most efficient combination to establish a biofilm. Cells that were no longer actively dividing and that lacked buoyancy displayed superior immobilization to cells that were actively dividing and buoyant. The immobilized cells did produce Microcystin but in much lower concentrations to cells grown in an airlift culture. Biofilms grown with a higher nitrate concentration, a lower biofilm thickness and a lower light intensity had a higher specific microcystin content, while biofilms with a higher nitrate concentration a lower light intensity and a longer growth period displayed the a greater toxin production per mm2 of membrane. Microcystin occurred at its highest concentration in cells just above the pore opening. The diffusion of nutrients occurred relatively quickly to the outside layers of the biofilm, with a true gradient being established laterally from these nutrient veins that were above the pores. Permeabilization of the cells proved unsuccessful, as cells that remained viable did not release the intracellular compound into the surrounding medium.

No description available.

The extraction of oil from microalgae was investigated. The study focused on the hydrothermal liquefaction of the microalgae Microcystis aeruginosa, Cyclotella meneghinia and Nitzschia pusilla. M. aeruginosa was collected from the Hartebeespoort dam, while C. meneghinia and N. pusilla were cultured in the laboratory. The experiments were conducted in a high pressure autoclave with an inert atmosphere. Sodium carbonate was studied as a potential catalyst. The hydrothermal liquefaction of M. aeruginosa, C. meneghinia and N. pusilla was carried out at various reaction temperatures and catalyst loads. For the liquefaction of M. aeruginosa the residence times were also varied. The reaction temperatures ranged from 260 to 340 °C, while the catalyst loads varied between 0 and 10 wt% Na2CO3. The residence time was varied between 15 and 45 minutes. The study showed that hydrothermal liquefaction of M. aeruginosa produced a maximum oil yield of 15.60 wt% at 300 °C, whereas the thermochemical liquefaction of C. meneghinia and N. pusilla produced maximum yields of 16.03 wt% and 15.33 wt%, respectively, at 340 °C. The residence time did not influence thermochemical liquefaction of the algae, while an increase in the catalyst load reduced the oil yield. The reaction conditions had no effect on the elemental composition or the calorific value of the thermochemical liquefaction oil. The calorific value of the hydrothermal liquefaction oils ranged from 28.57 to 35.90 MJ.kg -1 . Hydrothermal liquefaction of microalgae produced oil that can be used as substitute for coal in simple gasification processes. The study showed that microalgal blooms, such as the M. aeruginosa blooms of the Hartebeespoort dam, can be used for the extraction of oil through hydrothermal liquefaction. / Thesis (M.Ing. (Chemical Engineering))--North-West University, Potchefstroom Campus, 2010.

The extraction of oil from microalgae was investigated. The study focused on the hydrothermal liquefaction of the microalgae Microcystis aeruginosa, Cyclotella meneghinia and Nitzschia pusilla. M. aeruginosa was collected from the Hartebeespoort dam, while C. meneghinia and N. pusilla were cultured in the laboratory. The experiments were conducted in a high pressure autoclave with an inert atmosphere. Sodium carbonate was studied as a potential catalyst. The hydrothermal liquefaction of M. aeruginosa, C. meneghinia and N. pusilla was carried out at various reaction temperatures and catalyst loads. For the liquefaction of M. aeruginosa the residence times were also varied. The reaction temperatures ranged from 260 to 340 °C, while the catalyst loads varied between 0 and 10 wt% Na2CO3. The residence time was varied between 15 and 45 minutes. The study showed that hydrothermal liquefaction of M. aeruginosa produced a maximum oil yield of 15.60 wt% at 300 °C, whereas the thermochemical liquefaction of C. meneghinia and N. pusilla produced maximum yields of 16.03 wt% and 15.33 wt%, respectively, at 340 °C. The residence time did not influence thermochemical liquefaction of the algae, while an increase in the catalyst load reduced the oil yield. The reaction conditions had no effect on the elemental composition or the calorific value of the thermochemical liquefaction oil. The calorific value of the hydrothermal liquefaction oils ranged from 28.57 to 35.90 MJ.kg -1 . Hydrothermal liquefaction of microalgae produced oil that can be used as substitute for coal in simple gasification processes. The study showed that microalgal blooms, such as the M. aeruginosa blooms of the Hartebeespoort dam, can be used for the extraction of oil through hydrothermal liquefaction. / Thesis (M.Ing. (Chemical Engineering))--North-West University, Potchefstroom Campus, 2010.

Blooms of cyanobacteria have increased in occurrence in the past three decades and have been reported to cause severe problems for animals and humans, leading to death in extreme instances. The majority of poisonings that have taken place have been attributed to a hepatotoxin produced by the species Microcystis aeruginosa, namely microcystin. The appearance of a cyanobacterial bloom does not give any indication as to its toxicity and therefore, it is imperative that simple, yet sensitive, bioassays are developed to overcome this problem. This study was undertaken to evaluate the effects of microcystin-LR on the liver of Xenopus laevis both in vitro and in vivo. This animal provides an opportunity to study the long-term hepatotoxic effects of the toxin compared to in vitro studies performed with mice and rats. The use of the liver slice model system as a potential bioassay to study the effects of microcystin-LR on Xenopus laevis liver was evaluated. Liver slices were cultured in RPMI- 1640 culture medium for periods ranging from 30 hours to 10 days and the liver slices were exposed to toxin concentrations ranging from 1nM to 500nM. The use of frog liver slices to study the longer-term effects of low-dose exposure to microcystin-LR was evaluated by observing the ultrastructural changes within hepatocytes using transmission electron microscopy, the release of the enzymes alanine aminotransferase and lactate dehydrogenase into the surrounding culture medium, as well as using a 3-[4,5-dimethylthiazol-2yl]-2,5- diphenyl tetrazolium bromide assay to determine the viability of the liver slices in culture. The amount of lipid peroxidation in the liver slices after exposure to microcystin-LR was assessed using the Thiobarbituric Acid Test. Results showed the frog liver slice culture system to be an inadequate method to evaluate the hepatotoxic effects of microcystin-LR. An in vivo assessment of the effects of microcystin-LR on Xenopus laevis was carried out using a total of 9 frogs (3 groups of 3 frogs). Frogs received a single intraperitoneal dose of 120mg/kg of microcystin-LR and were sacrificed at 8 and 24 hours post exposure. Microcystin-LR caused no significant change in serum lactate dehydrogenase levels, hepatosomatic index (liver weight as a percentage body weight), glutathione peroxidase activity, glycogen or lipid peroxidation. There was, however, an increase in glutathione sii transferase activity in the liver. The presence of the toxin in the liver was confirmed by immunohistochemistry. This study suggests that Xenopus laevis has, in some way, adapted to detoxifying aquatic toxins in the environment.