A demanda crescente de água doce de boa qualidade são problemas atuais e mundiais, além do descaso com os dejetos lançados nos ambientes aquáticos que comprometem a qualidade dos recursos hídricos. Um dos parâmetros que atesta a potabilidade da água é a presença de cianobactérias e cianotoxinas. Cianobactérias são microrganismos procariontes aeróbicos fotoautróficos que sintetizam as cianotoxinas. Estes compostos podem ser classificados de acordo com seus mecanismos de ação em hepatotóxicos, neurotóxicos e dermatotóxicos. Por sua diversidade, representam diferentes riscos não só ao ecossistema e a outros organismos dos ambientes aquáticos, como também aos seres humanos. Esse projeto visou o isolamento e cultivo de cepas de cianobactérias produtoras de toxinas para a investigação da biossíntese desses compostos. Com este intuito, foram realizadas coletas de água em três reservatórios no estado de São Paulo e um no Paraná. Cepas de cianobactérais foram isoladas, identificadas e analisadas quanto à produção de toxinas. Uma cepa de Microcystis aeruginosa (LTPNA 02) produtora de microcistinas (MC-LR, MC-RR, MC-YR, MC-LF, MC-LW e desm-MC-LR e desm- MC-RR) foi escolhida para ser estudada frente diferentes condições de cultivo e ter o seu crescimento, produção de toxinas e expressão gênica estudados. Foram utilizados os meios de cultura já referidos na literatura: ASM-1 (N:P=1, 10 e 20), MLA (N:P=10), Bold 3N (N:P=16) e BG-11 (N:P=10 e 100). Para acompanhar o crescimento, dois métodos foram utilizados: contagem de células e espectrofotometria. As toxinas foram quantificadas por LC-MS - QTrap. A análise da expressão gênica foi realizada por reação de PCR em tempo real pelo método de quantificação relativa &#916;&#916;Ct. Foi observada diferença no crescimento da cepa estudada nos diferentes meios de cultivo empregados. A contagem das células permitiu a identificação das fases logarítmica e total de crescimento. Durante a fase logarítmica, três experimentos demonstraram diferenças estatísticas quando comparadas ao controle (p<0,05). Ao se avaliar o crescimento total, quatro experimentos foram menores (p<0,01). As leituras das absorvâncias e a contagem de células demonstraram alta correlação Para ambas as leituras em 680 nm e 750 nm o coeficiente de correlação (r) esteve entre 0,93 e 0,99. A quantificação das microcistinas (MC) foi realizada por LC-MS - QTrap. Foram quantificadas as variantes MC-LR, MR-RR e MC-YR. Apesar da relação toxina/célula ser distinto para cada experimento, não representou grande variação naqueles realizados com meio ASM-1 (N:P 1; 10 e 20), meio MLA (N:P=10) e BG11(N:P=10). O experimento realizado em Bold3N (N:P=16,6) apresentou menor concentração de toxina/célula e as variantes MC-LR e MC-YR não foram detectadas. Por outro lado, o experimento realizado em BG-11 (N:P=100) apresentou a maior relação toxina por célula. Estes resultados sugerem que o excesso de nitrato seja um fator estressante para o desenvolvimento e crescimento da cepa de M. aeruginosa avaliada e ao mesmo tempo um fator estimulante para a produção das toxinas analisadas. Os experimentos que avaliaram a expressão dos genes 16S e mcyB em relação ao gene da ficocianina (controle endógeno) foram realizados em meio ASM-1 (N:P=10 e 100) e BG 11 (N:P= 10 e 100). Os parâmetros anteriores, como crescimento e produção de toxinas também foram avaliados. Novamente foram encontradas diferenças entre as fases de crescimento e produção de toxina, porém a expressão dos genes avaliados não demonstrou variação significativa entre os experimentos. Porém ambos os genes avaliados demonstraram menor expressão nos experimentos condizidos em (N:P=100). / There is a great concern these days about potable and good quality water due to the increase of the population needs and also to the arising problems with contamination caused by anthropogenic sources. The presence of cyanobacteria and cyanotoxins are some parameters that attest water potability. Cyanobacteria are prokaryotic aerobic photoautotrophic microorganisms that may synthesize cyanotoxins. These compounds can be classified as hepatotoxic, neurotoxic and dermatotoxic according to their action mechanisms. Because of their diversity, they may represent different risks, not only to their ecosystem and other aquatic living organisms, but also to human beings. The aim of this project was the isolation and cultivation of cyanotoxin-producing cyanobacteria for further investigation on the biosynthesis of these compounds. Water samples from three different reservoirs in São Paulo state and one in Paraná state were collected in order to isolate cyanobacteria strains and accomplish their identification and to evaluate the toxin production. The Microcystis aeruginosa (LTPNA 02) microcystin producer strain (MCLR, MC-RR, MC-YR, MC-LF, MC-LW, desm-MC-LR and desm-MC-RR) was chosen to be grown in different cultivation conditions and later analyzed for its growth rate, toxin production and gene expression. All culture media used in this research were chosen according to the literature: ASM-1 (N:P=1, 10 and 20), MLA (N:P=10), Bold 3N (N:P=16) and BG-11 (N:P=10 and 100). To evaluate growth rate, two techniques were used: cell counting and absorbance determination in two different wavelengths (680 nm and 750 nm). Toxins were quantified by LC-MS in a hybrid triple-quadrupole instrument (Qtrap). Gene expression was assessed by real time PCR, using the &#916;&#916;Ct relative quantification method. Cell counting allowed total growth and logarithmic phase identification. During the last, three experiments showed statistical difference from control group (p<0,05). Four experiments resulted in a lower total growth rate (p<0,05). A high correlation between cell counting and absorbance levels was found for both wavelengths tested. Correlation coefficients (r) were from 0,93 to 0,99. Three microcystin variants (MC-LR, MR-RR e MC-YR) were quantified by LC-MS. The toxin content per cell was calculated and showed no statistc variation among those experiments performed on ASM-1 (N:P 1; 10 and 20), MLA (N:P=10) and BG-11 (N:P=10). The lowest toxin/cell concentration was found for Bold3N (N:P=16,6) medium, where MC-LR and MC-YR production was not detected. On the other hand, the experiment with BG-11 (N:P=100) medium showed the highest toxin/cell content. These results suggest that high levels of nitrate in the culture medium may be a stressing factor for the development and growth of the M. aeruginosa tested strain, as well as a disturbing factor for microcystin production. Gene expression experiments regarding 16S and mycB genes using the phycocyanin gene as endogen control were performed on ASM-1 (N:P=10 and 100) and BG 11 (N:P= 10 and 100) media, along with the evaluation of growth rate and toxin production. Differences between growth rates and toxin production were once more observed, however gene expression did not show a significant variation among experiments.

Microviridine sind ribosomal synthetisierte, cyanobakterielle Depsipeptide. Die genetische Basis der Microviridinproduktion ist ein Gencluster mit den Genen mdnABCDE. Zwei neuartige ATP-grasp-Ligasen, MdnB und MdnC, katalysieren die Bildung von Lacton- und Lactamringen durch die Einführung von zwei -Ester-und einer -Amidbindung. Die Prozessierung wird von einer bislang unbekannten Peptidase durchgeführt. Neben den filamentösen Nostoc und Planktothrix gehört die einzellige, blütenbildende Cyanobakteriengattung Microcystis zu den Microviridinproduzenten. Die inhibitorische Aktivität gegenüber Serinproteasen verleiht Microviridinen ökologische und pharmazeutische Relevanz. Im Rahmen dieser Arbeit wurde eine kleine Microviridin Expressionsplattform konstruiert. Ein neuartiges Microviridin Gencluster aus Microcystis aeruginosa Nies843 wurde heterolog in E. coli exprimiert, bioinformatisch analysiert und mutiert. Das hochkonservierte PFFARFL-Motif im Precursorpeptid MdnA wurde als Erkennungssequenz für die ATP-grasp Ligasen identifiziert. Manipulationen am C-Terminus des leader-Peptids führten zu einer Inhibierung der Aktivität von MdnB. Peptid-Protein-Interaktionen zwischen MdnA und den ATP-grasp Ligasen wurden untersucht. Der ABC-Transporter MdnE stabilisiert höchstwahrscheinlich einen Microviridin Biosynthesekomplex an der inneren Membran, wofür zwei mögliche Modelle vorgeschlagen werden. Punktmutationen in der Microviridin core-Sequenz offenbarten Flexibilität des Microviridin-Biosyntheseapparates für das peptide engineering. Es wurde eine Mutante konstruiert, deren inhibitorische Aktivität gegen Elastase um den Faktor 100 verbessert wurde. Durch die Konstruktion einer Precursoraustauschplattform konnten bisher kryptische Microviridine produziert werden. Diese Methode hat Potential für den Bau von Microviridinbibliotheken. Letztlich wird eine Hypothese zum Bindungsmechanismus von Microviridinen an Proteasen aufgestellt. / Microviridins are ribosomally synthesized cyanobacterial oligopeptides. These peptides comprise an unrivaled multicyclic cage-like structure, carrying two characteristic  ester and one amide bond, which are introduced by the two novel ATP-grasp ligases MdnB and MdnC. In addition to the filamentous species Nostoc and Planktothrix, the unicellular, bloom-forming cyanobacterium Microcystis aeruginosa Nies843 is one of the microviridin producer strains. The potent serine protease inhibitory activity contributes to both ecological and pharmacological relevance of microviridins. During this work, a small expression platform carrying the microviridin gene cassette mdnABCDE was established. Microviridins were heterologously expressed in E. coli and analyzed using bioinformatics and mutational analysis. The strictly conserved PFFARFL motif in the precursor peptide MdnA was identified and characterized as a binding sequence for the ATP-grasp ligases. Protein interactions of MdnA with B and C were studied. The ABC transporter MdnE was unveiled to be crucial for cyclization and processing of microviridins, probably stabilizing a putative microviridin maturation complex at the inner membrane. Two initial models for the peptide recognition and processing have been proposed. Point mutations in the microviridin core sequence showed some flexibility of the microviridin biosynthetic pathway to be used for peptide engineering. The exchange of a phenylalanine against a leucine in position 5 of the core region resulted in more than a 100-fold increased inhibitory activity against the attractive drug target elastase. The possibility to express cryptic microviridin precursor peptides in a precursor exchange platform showed the potential to create peptide libraries. Finally, a hypothesis about the binding mechanism of microviridins is presented.

Os efeitos de cianobactérias sobre organismos aquáticos planctônicos foram avaliados, visando caracterizar e quantificar as cianotoxinas e determinar a toxicidade de uma linhagem em cultura monoespecífica e de florações naturais de reservatórios do rio Tietê, SP. Assim, cultivou-se uma linhagem (NPLJ-4) de Microcystis aeruginosa, reconhecidamente tóxica, em meio ASM-1 a 25°C e fotoperíodo de 12h luz/12h escuro em câmara incubadora, avaliando-se sua toxicidade em diferentes estágios do crescimento populacional (meio e final da fase exponencial, fase estacionária e fase senescente), por meio de testes ecotoxicológicos com os organismos-teste Ceriodaphnia dubia e C. silvestrii. Esses testes foram realizados de acordo com normas padronizadas pela ABNT, sendo utilizados também para avaliar a toxicidade das florações naturais e a eficiência de diferentes processos de tratamento de água na remoção de células, microcistinas e subprodutos de cianobactérias. Os resultados indicaram aumento na concentração de microcistinas com o crescimento populacional da cianobactéria. Os extratos na fase estacionária tiveram menor toxicidade, enquanto nas demais fases causaram efeito tóxico agudo, resultando em valores de CE50; 48h de: 1,4 - 4,7 × \'10 POT.6\' cel/mL (meio fase exponencial), 1,6 - 8,7 × \'10 POT.6\' cel/mL (final fase exponencial), 7,5 - 14,1 × \'10 POT.6\' cel/mL (fase estacionária) e 1,9 - 4,6 × \'10 POT.6\' cel/mL (fase senescente) para C. dubia; e 1,9 - 5,4 × \'10 POT.6\' cel/mL (meio fase exponencial), 1,6 - 10,9 × \'10 POT.6\' cel/mL (final fase exponencial), 10,2 - 15,4 × \'10 POT.6\' cel/mL (fase estacionária) e 2,0 - 4,2 × \'10 POT.6\' cel/mL (fase senescente) para C. silvestrii. Células livres de Microcystis (M. aeruginosa, M. panniformis e M. protocystis) e Pseudanabaena mucicola foram as cianobactérias dominantes nas florações dos reservatórios de Barra Bonita e células livres de Microcystis (M. aeruginosa e M. panniformis), no de Promissão. A dominância das cianobactérias em ambos os reservatórios pode estar relacionada a períodos de estabilidade da coluna de água, razões N/P de 8 - 13 (Barra Bonita) e 19 - 20 (Promissão) na superfície, temperatura da água de 19 - 30°C (Barra Bonita) e 26 - 28°C (Promissão) e disponibilidade de nutrientes (0,05 - 0,26 mg/L de fósforo total para Barra Bonita e 0,01 - 0,05 mg/L P-total para Promissão), devido ao grau de trofia dos reservatórios. A água de ambos os reservatórios, coletada durante as florações, apresentou toxicidade aos dafinídeos, sendo que a água de Barra Bonita foi mais tóxica do que a de Promissão. Todos os extratos brutos de material oriundo das florações naturais apresentaram microcistinas (239 - 1647 µg/L para Barra Bonita e 192 - 1295 µg/L para Promissão) e causaram toxicidade aguda aos dafinídeos, com valores de CE50; 48h de: 87 - 282 mg/L (Barra Bonita) e 146 - 428 mg/L (Promissão) para C. dubia, e 98 - 546 mg/L (Barra Bonita) e 110 - 391 mg/L (Promissão) para C. silvestrii. Concentrações dos extratos a partir de 80 mg/L (Barra Bonita) e 100 mg/L (Promissão) afetaram adversamente a sobrevivência e a reprodução dos dafinídeos. Os resultados mostram riscos à biota natural e à saúde humana, bem como comprometimento dos usos múltiplos dos reservatórios, exigindo ações remediadoras e, sobretudo, preventivas para conter o processo de eutrofização. / The effects of cyanobacteria upon aquatic organisms were evaluated, aiming to characterize and quantify the toxins of both, a monospecific cyanobacterial culture and material from natural blooms occurring in the reservoirs of Tietê river, SP. The already known toxic strain NPLJ-4 of Microcystis aeruginosa was cultured in ASM-1 medium at 25 Celsius degrees and 12h light/12h dark in the incubator, in order to evaluate its toxicity at different stages of the culture growth by ecotoxicological tests using the cladocerans Ceriodaphnia dubia and C. silvestrii as test-organisms. These tests were carried out according to the procedures standardized by ABNT, in order to evaluate also the toxicity of natural blooms and the efficiency of different water treatment processes in removing cells, microcystins and by-products of cyanobacteria. The results obtained indicated an increase in the concentration of microcystins along the cyanobacterial culture growth. Extracts from the stationary phase of the culture were less toxic compared with those from the other phases which had acute toxicity and adversely affected cladoceran survival, resulting in EC50; 48h values of 1,4 - 4,7 × \'10 POT.6\' cells/mL (middle exponential phase), 1,6 - 8,7 × \'10 POT.6\' cells/mL (final exponential phase), 7,5 - 14,1 × \'10 POT.6\' cells/mL (stationary phase) and 1,9 - 4,6 × \'10 POT.6\' cells/mL (senescent phase) for C. dubia; and 1,9 - 5,4 × \'10 POT.6\' cells/mL (middle exponential phase), 1,6 - 10,9 × \'10 POT.6\' cells/mL (final exponential phase), 10,2 - 15,4 × \'10 POT.6\' cells/mL (stationary phase) and 2,0 - 4,2 × \'10 POT.6\' cells/mL (senescent phase) for C. silvestrii. Cells of Microcystis (Microcystis aeruginosa, M. panniformis and M. protocystis) and Pseudanabaena mucicola were cyanobacteria species dominant in the Barra Bonita reservoir and cells of Microcystis (M. aeruginosa and M. panniformis), in the Promissão reservoir. The dominance of cyanobacteria in both studied reservoirs was related to the stability of the water column, N/P ratios of 8 to 13 (Barra Bonita) and 19 to 20 (Promissão), high water temperatures (19 - 30°C for Barra Bonita and 26 - 28°C for Promissão) and high nutrient availability (0,05 - 0,26 mg/L total phosphorus for Barra Bonita, and 0,01 - 0,05 mg/L total P for Promissão) as a consequence of the trophic state of the reservoirs. The water from Barra Bonita reservoir during the cyanobacterial blooms was more toxic to daphnids than that from Promissão reservoir. Crude extracts from all cyanobacteria blooms tested presented microcystins (239 - 1647 µg/L for Barra Bonita and 192 - 1295 µg/L for Promissão) and caused acute toxicity to daphnids, resulting in EC50; 48h values of 87 - 282 mg/L (Barra Bonita) and 146 - 428 mg/L (Promissão) for C. dubia, and 98 - 546 mg/L (Barra Bonita) and 110 - 391 mg/L (Promissão) for C. silvestrii. Crude extracts concentrations above 80 mg/L to Barra Bonita and 100 mg/L to Promissão adversely affected the survival and reproduction of daphnids. The results obtained evidenced the risks to the natural biota and possibly to the human health, and can therefore jeopardize the multiple uses of the reservoirs. They reveal the urgent necessity for remedial action, particularly to slow down and to prevent eutrophication.

A demanda crescente de água doce de boa qualidade são problemas atuais e mundiais, além do descaso com os dejetos lançados nos ambientes aquáticos que comprometem a qualidade dos recursos hídricos. Um dos parâmetros que atesta a potabilidade da água é a presença de cianobactérias e cianotoxinas. Cianobactérias são microrganismos procariontes aeróbicos fotoautróficos que sintetizam as cianotoxinas. Estes compostos podem ser classificados de acordo com seus mecanismos de ação em hepatotóxicos, neurotóxicos e dermatotóxicos. Por sua diversidade, representam diferentes riscos não só ao ecossistema e a outros organismos dos ambientes aquáticos, como também aos seres humanos. Esse projeto visou o isolamento e cultivo de cepas de cianobactérias produtoras de toxinas para a investigação da biossíntese desses compostos. Com este intuito, foram realizadas coletas de água em três reservatórios no estado de São Paulo e um no Paraná. Cepas de cianobactérais foram isoladas, identificadas e analisadas quanto à produção de toxinas. Uma cepa de Microcystis aeruginosa (LTPNA 02) produtora de microcistinas (MC-LR, MC-RR, MC-YR, MC-LF, MC-LW e desm-MC-LR e desm- MC-RR) foi escolhida para ser estudada frente diferentes condições de cultivo e ter o seu crescimento, produção de toxinas e expressão gênica estudados. Foram utilizados os meios de cultura já referidos na literatura: ASM-1 (N:P=1, 10 e 20), MLA (N:P=10), Bold 3N (N:P=16) e BG-11 (N:P=10 e 100). Para acompanhar o crescimento, dois métodos foram utilizados: contagem de células e espectrofotometria. As toxinas foram quantificadas por LC-MS - QTrap. A análise da expressão gênica foi realizada por reação de PCR em tempo real pelo método de quantificação relativa &#916;&#916;Ct. Foi observada diferença no crescimento da cepa estudada nos diferentes meios de cultivo empregados. A contagem das células permitiu a identificação das fases logarítmica e total de crescimento. Durante a fase logarítmica, três experimentos demonstraram diferenças estatísticas quando comparadas ao controle (p<0,05). Ao se avaliar o crescimento total, quatro experimentos foram menores (p<0,01). As leituras das absorvâncias e a contagem de células demonstraram alta correlação Para ambas as leituras em 680 nm e 750 nm o coeficiente de correlação (r) esteve entre 0,93 e 0,99. A quantificação das microcistinas (MC) foi realizada por LC-MS - QTrap. Foram quantificadas as variantes MC-LR, MR-RR e MC-YR. Apesar da relação toxina/célula ser distinto para cada experimento, não representou grande variação naqueles realizados com meio ASM-1 (N:P 1; 10 e 20), meio MLA (N:P=10) e BG11(N:P=10). O experimento realizado em Bold3N (N:P=16,6) apresentou menor concentração de toxina/célula e as variantes MC-LR e MC-YR não foram detectadas. Por outro lado, o experimento realizado em BG-11 (N:P=100) apresentou a maior relação toxina por célula. Estes resultados sugerem que o excesso de nitrato seja um fator estressante para o desenvolvimento e crescimento da cepa de M. aeruginosa avaliada e ao mesmo tempo um fator estimulante para a produção das toxinas analisadas. Os experimentos que avaliaram a expressão dos genes 16S e mcyB em relação ao gene da ficocianina (controle endógeno) foram realizados em meio ASM-1 (N:P=10 e 100) e BG 11 (N:P= 10 e 100). Os parâmetros anteriores, como crescimento e produção de toxinas também foram avaliados. Novamente foram encontradas diferenças entre as fases de crescimento e produção de toxina, porém a expressão dos genes avaliados não demonstrou variação significativa entre os experimentos. Porém ambos os genes avaliados demonstraram menor expressão nos experimentos condizidos em (N:P=100). / There is a great concern these days about potable and good quality water due to the increase of the population needs and also to the arising problems with contamination caused by anthropogenic sources. The presence of cyanobacteria and cyanotoxins are some parameters that attest water potability. Cyanobacteria are prokaryotic aerobic photoautotrophic microorganisms that may synthesize cyanotoxins. These compounds can be classified as hepatotoxic, neurotoxic and dermatotoxic according to their action mechanisms. Because of their diversity, they may represent different risks, not only to their ecosystem and other aquatic living organisms, but also to human beings. The aim of this project was the isolation and cultivation of cyanotoxin-producing cyanobacteria for further investigation on the biosynthesis of these compounds. Water samples from three different reservoirs in São Paulo state and one in Paraná state were collected in order to isolate cyanobacteria strains and accomplish their identification and to evaluate the toxin production. The Microcystis aeruginosa (LTPNA 02) microcystin producer strain (MCLR, MC-RR, MC-YR, MC-LF, MC-LW, desm-MC-LR and desm-MC-RR) was chosen to be grown in different cultivation conditions and later analyzed for its growth rate, toxin production and gene expression. All culture media used in this research were chosen according to the literature: ASM-1 (N:P=1, 10 and 20), MLA (N:P=10), Bold 3N (N:P=16) and BG-11 (N:P=10 and 100). To evaluate growth rate, two techniques were used: cell counting and absorbance determination in two different wavelengths (680 nm and 750 nm). Toxins were quantified by LC-MS in a hybrid triple-quadrupole instrument (Qtrap). Gene expression was assessed by real time PCR, using the &#916;&#916;Ct relative quantification method. Cell counting allowed total growth and logarithmic phase identification. During the last, three experiments showed statistical difference from control group (p<0,05). Four experiments resulted in a lower total growth rate (p<0,05). A high correlation between cell counting and absorbance levels was found for both wavelengths tested. Correlation coefficients (r) were from 0,93 to 0,99. Three microcystin variants (MC-LR, MR-RR e MC-YR) were quantified by LC-MS. The toxin content per cell was calculated and showed no statistc variation among those experiments performed on ASM-1 (N:P 1; 10 and 20), MLA (N:P=10) and BG-11 (N:P=10). The lowest toxin/cell concentration was found for Bold3N (N:P=16,6) medium, where MC-LR and MC-YR production was not detected. On the other hand, the experiment with BG-11 (N:P=100) medium showed the highest toxin/cell content. These results suggest that high levels of nitrate in the culture medium may be a stressing factor for the development and growth of the M. aeruginosa tested strain, as well as a disturbing factor for microcystin production. Gene expression experiments regarding 16S and mycB genes using the phycocyanin gene as endogen control were performed on ASM-1 (N:P=10 and 100) and BG 11 (N:P= 10 and 100) media, along with the evaluation of growth rate and toxin production. Differences between growth rates and toxin production were once more observed, however gene expression did not show a significant variation among experiments.

A contínua entrada de metais pesados nos ambientes aquáticos constitui uma potencial ameaça aos ecossistemas naturais devido à ação tóxica direta em organismos aquáticos. Nos ambientes aquáticos, os organismos estão expostos a metais tanto dissolvidos na água como aqueles presentes na cadeia trófica. Um maior conhecimento sobre o papel do alimento como rota adicional de exposição a metais ou como um possível retentor de sua toxicidade para invertebrados aquáticos é necessário. Considerando-se a importância dos metais na contaminação ambiental, bem como a necessidade de melhor entendimento das interações desses elementos nos sistemas aquáticos, o presente estudo visou a avaliar a sensibilidade de espécies fitoplanctônicas (Selenastrum capricornutum e Microcystis aeruginosa) e de espécies zooplanctônicas (Daphnia similis e Ceriodaphnia dubia) aos metais cádmio e cromo e os efeitos tóxicos desses elementos em tais organismos. O crescimento celular, a concentração de clorofila, o biovolume e o peso seco das algas foram analisados quando as espécies algais foram expostas aos metais por meio de testes de toxicidade aguda. Testes de toxicidade aguda e crônica com o zooplâncton aos metais também foram realizados e o efeito de diferentes densidades algais (alta, média e baixa) sobre a toxicidade dos metais aos cladóceros foi avaliado. Além disso, algas foram expostas aos metais, oferecidas como alimento a C. dubia e os efeitos tóxicos crônicos foram investigados. Os resultados demonstraram que S. capricornutum foi mais sensível ao cádmio e M. aeruginosa foi mais sensível ao cromo, sendo essa diferença relacionada à capacidade das algas para reter os metais. Com o aumento de ambos os metais, houve uma diminuição na densidade celular, na taxa de crescimento, na clorofila e no peso seco das algas. A presença de diferentes densidades de S. capricornutum não alterou significativamente o valor da CE(I)50; 48h aos metais para D. similis, mas a elevada densidade de M. aeruginosa reduziu a toxicidade do cádmio para o dafinídeo. A alta densidade de alimento (106 céls/mL) influenciou negativamente a reprodução e a sobrevivência de C. dubia quando exposta a concentrações subletais dos metais em solução. Alimento exposto à metais, quando fornecido em alta e em média densidade, também afetou a sobrevivência e a reprodução dos organismos-teste. Apesar de a água ser uma importante rota de exposição aos metais para os organismos aquáticos, o alimento deve ser considerado uma via de contaminação adicional. / The continuous input of heavy metals into aquatic systems constitutes a potential threat to natural ecosystems because of the direct toxic action on aquatic organisms. In aquatic ecosystems, the organisms are exposed to dissolved metals in the water as to metals presents in the food chain. A larger knowledge about paper of food as an additional route of exposure to metals or as possible retainer of its toxicity for aquatic organisms is necessary. Considering the importance of metals in contamination of aquatic ecosystems and the need of better understanding of the interactions of those elements in the aquatic ecosystems as well, the aim of this study was to evaluate the sensibility of phytoplankton species (Selenastrum capricornutum and Microcystis aeruginosa) and zooplankton species (Daphnia similis and Ceriodaphnia dubia) to cadmium and chromium metals and the toxic effects of those elements in such organisms. Analysed of cellular growth, chlorophyll concentration, biovolume and dry weight of the algae were carried out when algal species were exposed to metals. Acute and chronic toxicity tests with the zooplankton were also accomplished and the effect of different algal densities (high, middle and low) on toxicity of metals to cladoceran was evaluated. Alga S. capricornutum was exposed to metals, supplied as food to C. dubia and chronic toxicity effects were investigated. The results demonstrated that S. capricornutum was more sensitive to cadmium and M. aeruginosa was more sensitive to chromium, this difference was related the capacity of the algae to retain metals. Cell density, growth rate, chlorophyll and dry weight of the algae were reduced with increase of both metals. The presence of different density of S. capricornutum does not alter the value of EC50 to metals for D. similis, but high density of M. aeruginosa reduced the toxicity of cadmium for daphnid. High food density (106 cells/mL) influenced negatively on reproduction and survival of C. dubia when this organism was exposed to sublethal concentrations of metals in solution. Food exposed to metals, when it was supplied at high and middle density, also affected survival and reproduction of the test organisms. Although the water to be considered a important route of exposure of metals for aquatic organisms, the food should be considered an additional source of toxicity.

Sowohl in Süßwasserseen als auch in marinen Gewässern kommt es immer wieder zu Massenentwicklungen von Cyanobakterien, sogenannten “Blüten”. In Seen werden diese oftmals von Cyanobakterien der Gattung Microcystis dominiert, deren Arten häufig Toxine bilden. Die verbreitesten dieser Toxine sind die leberschädigen Microcystine, die eine Klasse nichtribosomal synthetisierter Peptide darstellen. Nachdem die toxische Wirkung der Microcystine bisher als deren Hauptfunktion angesehen wurde, deuten neuere Forschungsergebnisse darauf hin, dass Microcystine eine andere Primärfunktion für die Produzenten besitzen. Im Rahmen dieser Studie wurde Microvirin (Mvn), ein putatives Lektin aus Microcystis aeruginosa PCC 7806, von dem angnommen wurde, dass es funktional mit Microcystin assoziiert ist, charakterisiert. Zunächst konnte gezeigt werden, dass Mvn tatsächlich zuckerbindende Aktivität besitzt und spezifisch Mannan, ein Oligosaccharid aus Mannoseuntereinheiten, erkennt. Bindestudien zeigten, dass Zucker dieses Typs auf der Zelloberfläche von M. aeruginosa PCC 7806 lokalisiert sind und eine Bindestelle für das sekretierte Mvn darstellen. Mit Hilfe fluoreszenzmikroskopiebasierender Methoden wurde gezeigt, dass sowohl Mvn als auch das korrespondierende Mannanoligosaccharid stammspezifisch sind. Weiterhin konnte durch PCR gezeigt werden, dass das mvn-Gen in allen getesteten Microcystis-Stämmen vorkommt, die auch Gene für die Microcystinbiosynthese besitzen. Eine direkte Interaktion von Microcystin und Mvn konnte in vitro bestätigt werden. Microcystin bindet dabei über seinen N-Methyl-Dehydroalaninrest kovalent an die reduzierten Cysteinreste des Proteins. Ein Einfluss auf die Oligomerisierung des Proteins wurde festgestellt. Microcystin bindet an Cysteinreste von Proteinen, und es konnte gezeigt werden, dass dies besonders unter oxidativen Stressbedingungen geschieht. Die Daten liefern somit weitere Indizien für eine Rolle von Microcystin in der Stressadaptation. / Cyanobacteria frequently appear as so-called “water-blooms” during summer months. Cyanobacteria of the genus Microcystis, whose species often dominate freshwater lakes, produce toxins that represent a potential threat for humans and animals. The most prominent toxins are the non-ribosomally synthesised hepatotoxic microcystins. Toxicity has been considered the main function of these peptides, but recent studies propose different primary functions of microcystins for their producers. The involvement of microcystins in the response to oxidative stress was proposed recently. Within this study the putative lectin microvirin (Mvn), which was suggested to be functionally related to microcystin, was characterised. Initially it was shown that Mvn does indeed possess a carbohydrate binding activity, and specificity for mannan, an oligosaccharide made of mannose subunits, was proven. Binding studies using fluorescence-labelled Mvn and antibodies identified carbohydrates of this type at the cell surface of M. aeruginosa being a binding site for the secreted Mvn. Fluorescence microscopy techniques were employed to show that Mvn as well as the corresponding mannan oligosaccharide are strain-specific. Additionally it was shown by PCR that the mvn gene is present in all tested Microcystis strains possessing microcystin biosynthesis genes. A direct interaction of microcystin and Mvn was confirmed in vitro. Microcystin covalently binds to the reduced cysteine residues of the protein via its N-methyl-dehydroalanine moiety. An impact on the oligomerisation state of Mvn was observed. Microcystin seems to bind cysteine residues in an unspecific manner in vivo, and it was shown that this occurs especially under conditions of oxidative stress such as iron depletion and exposition to high light. Hence, the data provide further evidence for an involvement of microcystins in stress adaptation.

Cyanobakterien bilden zahlreiche bioaktive Substanzen mit zum Teil humantoxischer Relevanz. Nicht selten spielen dabei zyklische Peptide, zu denen unter anderem die Microcystine (MCYST) gehören, eine Schlüsselrolle. MCYST werden u.a. von Microcystis KÜTZING EX LEMMERMANN 1907 gebildet. Erkenntnisse zur ökophysiologischen Funktion der MCYST, die zweifelsfrei bei den Produzenten selbst zu suchen ist, liegen bisher kaum vor. Mit Hilfe von Freilanduntersuchungen sollten im Rahmen der vorliegenden Arbeit Kenntnisse zu einer möglichen ökologischen Funktion der MCYST erweitert und vertieft werden. Grundlage stellte dabei die Phänologie von Microcystis als einer der bedeutendsten limnischen MCYST-Produzenten dar. Microcystis zeigt im Freiland einen charakteristischen annuellen Lebenszyklus mit benthisch-pelagischer Kopplung. Ziel der vorliegenden Arbeit war es, die phänologischen Phasen des Lebenszyklus von Microcystis im Freiland zu differenzieren sowie die Dynamik der MCYST während dieser Phasen kompartimentübergreifend gesamtheitlich zu erfassen. Über eine MCYST-Massenbilanzierung sollen anschließend die dem annuellen Zyklus zugrundeliegenden Teilprozesse quantifiziert und zusammengeführt werden. Vordergründiges Anliegen war es, Phasen einzugrenzen, bei denen MCYST möglicherweise eine ökophysiologische Funktion haben könnte. Der annuelle Lebenszyklus von Microcystis wurde anhand von Biomasseänderungen am Sediment und im Pelagial der TS Quitzdorf in die phänologischen Phasen Überwinterung, Reinvasion, pelagisches Wachstum und Sedimentation unterteilt: Intakte, im Herbst aus dem Freiwasser aussedimentierte, Microcystis-Kolonien überwintern am Sediment und steigen im Frühjahr und Frühsommer zurück ins Freiwasser auf. Dort erfolgt der Wachstumsprozess, dem sich im darauffolgenden Herbst erneut ein Zusammenbruch und die Sedimentation der Freiwassergemeinschaft anschließt. Die benthisch-pelagische Kopplung wirkt dabei als interannuelles Bindeglied. Zwischen dem annuellen Lebenszyklus von Microcystis und der MCYST-Dynamik wurde eine enge Bindung nachgewiesen: Änderungen der absoluten MCYST-Konzentrationen während der Übergangsphasen Aufstieg (Frühjahr) und Sedimentation (Herbst) zeigen, dass MCYST mit den aufsteigenden bzw. aussedimentierenden Microcystis-Kolonien aus dem bzw. in das Sediment ‚transportiert’ werden. Ausschließlich während der pelagischen Phase, die sich dem Reinvasionsprozess anschließt, kommt es in Abhängigkeit vom Wachstum der Produzenten und deren Sukzession zur Neubildung von MCYST. Während den Wintermonaten wurden MCYST am Sediment intrazellulär ‚konserviert’. Der Verlauf der pelagischen MCYST-Konzentration wurde mit Hilfe eines Wachstumsmodells nachgebildet. In dieses Modell wurde die genetische Variabilität der MCYST-Produzenten sowie eine mögliche physiologische Steuerung der MCYST-Synthese über die Verfügbarkeit des anorganischen Kohlenstoffs integriert. Der prinzipielle Verlauf zeigte dabei weitestgehend Koinzidenz zwischen den real gemessenen und den simulierten MCYST-Konzentrationswerten. Abweichungen zwischen beiden konnten mit Hilfe des gesamtheitlich kompartimentübergreifenden MCYST-Bilanzierungsansatzes – in erster Linie über benthisch-pelagische Kopplungsprozesse – plausibel erklärt werden. Der Habitatwechsel ist für Microcystis prinzipiell mit Verlusten (Seneszenz/Lyse oder möglicherweise Apoptose) verbunden, sowohl für MCYST-Produzenten und Nichtproduzenten. Die auffallende Stabilität der benthischen MCYST-Zellquote während der Überwinterung gibt Grund zur Annahme, dass eine Funktion von MCYST am/im Sediment eher unwahrscheinlich ist. Da MCYST über derart lange Zeiträume am Sediment intrazellulär ‚konserviert’ werden, ist eine Bedeutung der MCYST während der Reinvasionsphase und in der frühen pelagischen Phase nicht auszuschließen. Im Speziellen wurde eine mögliche ökologische Funktion von MCYST in Zusammenhang mit der Variation der Koloniegröße bzw. dem epiphytischen Bewuchs von Microcystis-Kolonien mit Pseudanabaena mucicola geprüft: Aus dem Zusammenhang zwischen extra-/intrazellulärer MCYST-Konzentration und der Microcystis-Koloniegrößenverteilung waren keine konsistenten Schlussfolgerungen abzuleiten, welche auf eine Steuerung der Koloniebildung durch MCYST deuten. Vor dem Hintergrund, dass MCYST keinen nachweislich allelopathischen Effekt auf den Epibionten Pseudanabaena mucicola ausüben, wurde postuliert, dass zwischen dem beobachteten epiphytischen Besiedlungs-/Verteilungsmuster und der MCYST-Produktion ein indirekter Zusammenhang besteht, welcher die zeitweise Einnischung von Pseudanabaena mucicola auf Microcystis-Kolonien ermöglicht. Die Ergebnisse der vorliegenden Untersuchung lassen weder unmittelbar noch mittelbar eine Variabilität der ökophysiologischen Bedeutung von MCYST, die im Zusammenhang mit der raum-zeitlichen Verteilung potentieller Produzenten steht, erkennen. Eine divergierende Funktion der MCYST auf intra- bzw. extrazellulärer Ebene kann nicht zwingend ausgeschlossen werden. Die Mehrzahl der aus der MCYST-Phänologie und MCYST-Bilanzierung abzuleitenden Schlussfolgerungen deutet allerdings eher auf eine Funktion auf (intra-)zellulärer Ebene hin, wie etwa die Effizienzsteigerung des Kohlenstoffmetabolismus (d.h. der intrazellulä-ren Akkumulation anorganischen Kohlenstoffs) während der pelagischen (Wachstums-)Phase der Produzenten.

Les proliférations de cyanobactéries présentent en général d'importantes variations spatiales et temporelles de leur abondance cellulaire et de leur potentiel toxique, ce qui rend très difficile la surveillance de ces microorganismes et l'estimation des risques sanitaires associés à ces événements. Dans ce cadre général, le premier objectif de ma thèse a été de tester différentes stratégies d'échantillonnage pour suivre au mieux l'évolution des abondances cellulaires lors des proliférations de cyanobactéries. Par un échantillonnage à haute fréquence (six points tous les deux jours) réalisé sur un étang piscicole du Forez, nous avons montré que les deux espèces qui proliféraient dans cet écosystème (Microcystis aeruginosa et Aphanizomenon flos-aquae) présentaient des dynamiques spatiales et temporelles de leur abondance cellulaire très contrastées, ne permettant pas de définir une stratégie d'échantillonnage optimale commune aux deux espèces. Si pour ces deux espèces, trois points d'échantillonnage sont au minimum nécessaires pour bien prendre en compte l'hétérogénéité dans la distribution spatiale des cellules, les fréquences d'échantillonnage optimales sont en revanche très variables, allant d'un échantillonnage mensuel ou bimensuel pour Microcystis à un échantillonnage au minimum hebdomadaire pour Aphanizomenon. Le second objectif de ma thèse a été de m'appuyer sur cette stratégie d'échantillonnage à haute fréquence, pour mieux comprendre le développement de la prolifération de M. aeruginosa. Pour ce faire, nous avons estimé les changements spatiaux et temporels intervenant dans la composition génotypique de la population de cyanobactéries (par utilisation de la SSCP sur l'ITS 16S-23S) et dans sa toxicité potentielle (par le dosage des microcystines et l'évaluation de la proportion de cellules possédant le gène mcyB, un des gènes de synthèse de la microcystine). La répartition spatiale de la composition génotypique s'est révélée homogène à l'échelle de l'étang. Il est également apparu qu'au cours de la phase de croissance de la population, cette composition génotypique subissait de nombreux changements à courte échelle temporelle puis restait stable pendant plusieurs semaines lorsque le maximum d'abondance était atteint. Au niveau du potentiel toxique, la proportion de cellules possédant le gène mcyB est resté proche de 60 % durant toute la prolifération avec toutefois, des variations plus importantes pendant le développement du bloom. Aucune relation n'a pu être établie entre les variations observées dans la composition génotypique de la population et celles dans les proportions de cellules toxiques. De même, aucun lien n'a pu être établi entre les variations de diverses variables environnementales (nutriments, température, pluie) et celles de l'abondance cellulaire, de la toxicité potentielle et de la composition génotypique de la population de Microcystis. L'ensemble de ces résultats suggère que d'autres facteurs et processus interviennent probablement dans ces variations et qu'il existe sans doute des interactions très complexes entre toutes ces variables. Enfin, le troisième objectif de ma thèse a été de comparer la composition génotypique et le potentiel toxique de plusieurs populations de Microcystis plus ou moins connectées entre elles. Cette comparaison a permis de montrer l'importance fondamentale des facteurs et processus environnementaux locaux, dans la mise en place et le développement de ces évènements. / Generally, cyanobacteria proliferations show great spatial and temporal variations in their cell abundances and potential toxicity, which makes it difficult to control the development of these microorganisms and to predict the health risks associated with these events. Within this scope, the first goal of my PhD thesis was to test different sampling strategies to guarantee the best monitoring of the cell abundances during cyanobacteria proliferations. We made a high frequency sampling (six points every other day) in a shallow lake located in Forez and we evidenced that the two blooming-species (Microcystis aeruginosa and Aphanizomenon flos-aquae) showed strongly contrasted spatial and temporal patterns of their cell abundances precluding having a common optimal sampling strategy for both species. Even if three sampling points were enough to take into account the spatial heterogeneity of Microcystis and Aphanizomenon cells, a monthly or two-monthly sampling was sufficient for Microcystis whereas a weekly sampling was necessary for Aphanizomenon. The second goal was to gain a better understanding of Microcystis proliferation development. To achieve this aim, we estimated spatial and temporal changes in the genotypic composition (using the SSCP method in the 16S-23S ITS) and in the potential toxicity (by measuring the microcystin concentration and proportion of mcyB+ cells). We obtained a homogeneous spatial repartition of the genotypic composition. Moreover, during the growth phase, there were many rapid changes in the genotypic composition whereas this composition remained stable for several weeks where the maximum cell abundance was reached. As for potential toxicity, the proportion of mcyB+ cells remains at around 60 % during the proliferation but we observed higher variations during the growth phase. No relation was found between the variations of the genotypic composition and proportion of toxic cells on the one hand and the variations of several environmental factors (nutrients, temperature, rain) on the other hand, suggesting that other factors may be involved in these variations and that many complex interactions occur between these factors. Finally, the third goal of my PhD thesis was to compare the genotypic composition and the potential toxicity of different Microcystis populations, which were more or less interconnected. This comparison evidenced the great importance of local environmental factors and processes in the beginning and development of these events.

Os efeitos de cianobactérias sobre organismos aquáticos planctônicos foram avaliados, visando caracterizar e quantificar as cianotoxinas e determinar a toxicidade de uma linhagem em cultura monoespecífica e de florações naturais de reservatórios do rio Tietê, SP. Assim, cultivou-se uma linhagem (NPLJ-4) de Microcystis aeruginosa, reconhecidamente tóxica, em meio ASM-1 a 25°C e fotoperíodo de 12h luz/12h escuro em câmara incubadora, avaliando-se sua toxicidade em diferentes estágios do crescimento populacional (meio e final da fase exponencial, fase estacionária e fase senescente), por meio de testes ecotoxicológicos com os organismos-teste Ceriodaphnia dubia e C. silvestrii. Esses testes foram realizados de acordo com normas padronizadas pela ABNT, sendo utilizados também para avaliar a toxicidade das florações naturais e a eficiência de diferentes processos de tratamento de água na remoção de células, microcistinas e subprodutos de cianobactérias. Os resultados indicaram aumento na concentração de microcistinas com o crescimento populacional da cianobactéria. Os extratos na fase estacionária tiveram menor toxicidade, enquanto nas demais fases causaram efeito tóxico agudo, resultando em valores de CE50; 48h de: 1,4 - 4,7 × \'10 POT.6\' cel/mL (meio fase exponencial), 1,6 - 8,7 × \'10 POT.6\' cel/mL (final fase exponencial), 7,5 - 14,1 × \'10 POT.6\' cel/mL (fase estacionária) e 1,9 - 4,6 × \'10 POT.6\' cel/mL (fase senescente) para C. dubia; e 1,9 - 5,4 × \'10 POT.6\' cel/mL (meio fase exponencial), 1,6 - 10,9 × \'10 POT.6\' cel/mL (final fase exponencial), 10,2 - 15,4 × \'10 POT.6\' cel/mL (fase estacionária) e 2,0 - 4,2 × \'10 POT.6\' cel/mL (fase senescente) para C. silvestrii. Células livres de Microcystis (M. aeruginosa, M. panniformis e M. protocystis) e Pseudanabaena mucicola foram as cianobactérias dominantes nas florações dos reservatórios de Barra Bonita e células livres de Microcystis (M. aeruginosa e M. panniformis), no de Promissão. A dominância das cianobactérias em ambos os reservatórios pode estar relacionada a períodos de estabilidade da coluna de água, razões N/P de 8 - 13 (Barra Bonita) e 19 - 20 (Promissão) na superfície, temperatura da água de 19 - 30°C (Barra Bonita) e 26 - 28°C (Promissão) e disponibilidade de nutrientes (0,05 - 0,26 mg/L de fósforo total para Barra Bonita e 0,01 - 0,05 mg/L P-total para Promissão), devido ao grau de trofia dos reservatórios. A água de ambos os reservatórios, coletada durante as florações, apresentou toxicidade aos dafinídeos, sendo que a água de Barra Bonita foi mais tóxica do que a de Promissão. Todos os extratos brutos de material oriundo das florações naturais apresentaram microcistinas (239 - 1647 µg/L para Barra Bonita e 192 - 1295 µg/L para Promissão) e causaram toxicidade aguda aos dafinídeos, com valores de CE50; 48h de: 87 - 282 mg/L (Barra Bonita) e 146 - 428 mg/L (Promissão) para C. dubia, e 98 - 546 mg/L (Barra Bonita) e 110 - 391 mg/L (Promissão) para C. silvestrii. Concentrações dos extratos a partir de 80 mg/L (Barra Bonita) e 100 mg/L (Promissão) afetaram adversamente a sobrevivência e a reprodução dos dafinídeos. Os resultados mostram riscos à biota natural e à saúde humana, bem como comprometimento dos usos múltiplos dos reservatórios, exigindo ações remediadoras e, sobretudo, preventivas para conter o processo de eutrofização. / The effects of cyanobacteria upon aquatic organisms were evaluated, aiming to characterize and quantify the toxins of both, a monospecific cyanobacterial culture and material from natural blooms occurring in the reservoirs of Tietê river, SP. The already known toxic strain NPLJ-4 of Microcystis aeruginosa was cultured in ASM-1 medium at 25 Celsius degrees and 12h light/12h dark in the incubator, in order to evaluate its toxicity at different stages of the culture growth by ecotoxicological tests using the cladocerans Ceriodaphnia dubia and C. silvestrii as test-organisms. These tests were carried out according to the procedures standardized by ABNT, in order to evaluate also the toxicity of natural blooms and the efficiency of different water treatment processes in removing cells, microcystins and by-products of cyanobacteria. The results obtained indicated an increase in the concentration of microcystins along the cyanobacterial culture growth. Extracts from the stationary phase of the culture were less toxic compared with those from the other phases which had acute toxicity and adversely affected cladoceran survival, resulting in EC50; 48h values of 1,4 - 4,7 × \'10 POT.6\' cells/mL (middle exponential phase), 1,6 - 8,7 × \'10 POT.6\' cells/mL (final exponential phase), 7,5 - 14,1 × \'10 POT.6\' cells/mL (stationary phase) and 1,9 - 4,6 × \'10 POT.6\' cells/mL (senescent phase) for C. dubia; and 1,9 - 5,4 × \'10 POT.6\' cells/mL (middle exponential phase), 1,6 - 10,9 × \'10 POT.6\' cells/mL (final exponential phase), 10,2 - 15,4 × \'10 POT.6\' cells/mL (stationary phase) and 2,0 - 4,2 × \'10 POT.6\' cells/mL (senescent phase) for C. silvestrii. Cells of Microcystis (Microcystis aeruginosa, M. panniformis and M. protocystis) and Pseudanabaena mucicola were cyanobacteria species dominant in the Barra Bonita reservoir and cells of Microcystis (M. aeruginosa and M. panniformis), in the Promissão reservoir. The dominance of cyanobacteria in both studied reservoirs was related to the stability of the water column, N/P ratios of 8 to 13 (Barra Bonita) and 19 to 20 (Promissão), high water temperatures (19 - 30°C for Barra Bonita and 26 - 28°C for Promissão) and high nutrient availability (0,05 - 0,26 mg/L total phosphorus for Barra Bonita, and 0,01 - 0,05 mg/L total P for Promissão) as a consequence of the trophic state of the reservoirs. The water from Barra Bonita reservoir during the cyanobacterial blooms was more toxic to daphnids than that from Promissão reservoir. Crude extracts from all cyanobacteria blooms tested presented microcystins (239 - 1647 µg/L for Barra Bonita and 192 - 1295 µg/L for Promissão) and caused acute toxicity to daphnids, resulting in EC50; 48h values of 87 - 282 mg/L (Barra Bonita) and 146 - 428 mg/L (Promissão) for C. dubia, and 98 - 546 mg/L (Barra Bonita) and 110 - 391 mg/L (Promissão) for C. silvestrii. Crude extracts concentrations above 80 mg/L to Barra Bonita and 100 mg/L to Promissão adversely affected the survival and reproduction of daphnids. The results obtained evidenced the risks to the natural biota and possibly to the human health, and can therefore jeopardize the multiple uses of the reservoirs. They reveal the urgent necessity for remedial action, particularly to slow down and to prevent eutrophication.

Made available in DSpace on 2014-12-17T15:03:31Z (GMT). No. of bitstreams: 1 LucianaCM_DISSERT.pdf: 1660460 bytes, checksum: dab339dbdc22796b2b9d04615f1dc630 (MD5) Previous issue date: 2013-04-17 / Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico / The potentially toxic cyanobacterial blooms in water bodies are spread across the globe, resulting in the loss of water quality and adverse effects on human health. In arid and semiarid regions, the hydrologic regime characterized by an annual cycle of drought and rain, change the volume and the retention time of the reservoir. Such changes affect the limnological characteristics and causing changes in composition and biomass community of cyanobacteria. The reservoir Cruzeta (Zmax = 8.7 m) is a eutrophic water supply source located in the semiarid tropical (Northeast Brazil). Raised the hypothesis that the hydrological regime of semi-arid tropical is a determining factor in the availability of resources in eutrophic water sources, which influences the composition of dominant species of cyanobacteria. The aim of this study was to analyze the changes in biomass and species composition of cyanobacteria for two annual hydrological cycles and evaluate factors drivers. The study was divided into five distinct periods (dry 2010, rain 2011, dry 2011, rain 2012, dry 2012). The dominant group found in all periods was Cyanobacteria (99% of total biomass), which contributed to the low diversity. The filamentous species Cylindrospermopsis raciborskii was present at both points in almost every study. The colonial species Microcystis panniformis and Sphaerocavum brasiliensis dominated only in periods with lower volumes of water. The diatoms contribute more to the biomass during the period of severe drought. The point near the dam (P1) had phytoplankton biomass larger than the point near the tributary (P2). The dominant species of colonial cyanobacteria lasted until the overflow in P1, and P2 this dominance was until the first rains. The redundancy analysis indicated that physical factors such as light availability and water level were the main factors driving the seasonal succession of phytoplankton. The composition of phytoplankton in spring was alternated by species of filamentous cyanobacteria in conditions of poor stability of the water column, such as Cylindrospermopsis raciborskii, and colonial species under conditions of high stability of the water column, such as Microcystis panniformis and Sphaerocavum brasiliensis. The extremes of torrential rains and severe droughts, governed by the hydrological regime of the semi-arid region led to the availability of resources in the watershed, directing the spatial and temporal dynamics of phytoplankton in the reservoir Cruzeta / As flora??es de cianobact?rias potencialmente t?xicas est?o disseminadas em corpos aqu?ticos por todo o globo, resultando na perda da qualidade da ?gua e efeitos negativos para a sa?de humana. Em regi?es ?ridas e semi?ridas, o regime hidrol?gico, caracterizado por um ciclo anual de seca e chuva, altera o volume e o tempo de reten??o dos reservat?rios. Tais altera??es afetam as caracter?sticas limnol?gicas e, consequentemente, ocassiona mudan?as na composi??o e biomassa da comunidade de cianobact?rias. O reservat?rio Cruzeta (Zmax = 8,7 m) ? um manancial de abastecimento eutr?fico, localizado no semi?rido tropical (Nordeste, Brasil). Levantou-se a hip?tese de que o regime hidrol?gico do semi?rido tropical ? um fator determinante na disponibilidade de recursos em mananciais eutrofizados, o que influencia na composi??o e domin?ncia de esp?cies de cianobact?rias. O objetivo deste trabalho foi analisar as mudan?as na biomassa e na composi??o de esp?cies de cianobact?rias durante dois ciclos hidrol?gicos anuais e avaliar fatores direcionadores. O estudo foi dividido em 5 per?odos distintos (seca 2010, chuva 2011, seca 2011, chuva 2012, seca 2012). O grupo dominante encontrado em todos os per?odos foi de Cianobact?rias (99% da biomassa total), o que contribuiu para a baixa diversidade. A esp?cie filamentosa Cylindrospermopsis raciborskii esteve presente em ambos os pontos em quase todo o estudo. As esp?cies coloniais Microcystis panniformis e Sphaerocavum brasiliensis dominaram somente nos per?odos com menores volumes de ?gua. As diatom?ceas contribu?ram mais com a biomassa durante o per?odo de seca severa. O ponto pr?ximo a barragem (P1) apresentou biomassa fitoplant?nica maior que o ponto pr?ximo ao tribut?rio (P2). A domin?ncia das esp?cies coloniais de cianobact?rias se estendeu at? o extravazamento do reservat?rio no P1. No P2, esta domin?ncia ocorreu at? as primeiras chuvas. A an?lise de redund?ncia indicou que vari?veis f?sicas, tais como disponibilidade de luz e o volume de ?gua, foram os principais fatores de condu??o da sucess?o sazonal do fitopl?ncton. A composi??o do fitopl?ncton no manancial foi alternada por esp?cies de cianobact?rias filamentosas nas condi??es de pouca estabilidade da coluna d??gua, como a Cylindrospermopsis raciborskii, e por esp?cies coloniais nas condi??es de elevada estabilidade da coluna d??gua, como Microcystis panniformis e Sphaerocavum brasiliensis. Os eventos extremos de chuvas torrenciais e secas severas, regidos pelo regime hidrol?gico da regi?o do semi?rido determinaram a disponibilidade de recursos no manancial, direcionando a din?mica temporal e espacial do fitopl?ncton no reservat?rio Cruzeta