Dados da dissertação de mestrado
Author: Cesar Macedo Lima Filho
Title: Evaluation of the capacity of phytoremediation by Eichhornia crassipes (Mart.) Solms (Pontederiaceae)
Year: 2019 Full text (in Portuguese)
Abstract:
Eutrophication of water bodies is a worldwide problem. Eichhornia crassipes is a floating macrophyte with ability to remove nutrients from eutrophic aquatic environments and can be used for phytoremediation. This study was performed in the Barra do Braúna hydroelectric plant (MG, Brazil). A system consisting of 5 cascading mesocosms at decreasing heights was constructed (total volume on 15,000 L). The system was fed with water from the reservoir and sustained a flow of q = 89 L/h. To simulate a hypereutrophic state, NPK 10-10-10 was added to the system to increase the concentration of PO4 3- to 400 μg/L. To evaluate the phytoremediation capacity of E. crassipes, 30 plants were placed in each mesocosm. Two experiments were performed: one in the rainy season, summer (February - March) and the other in the dry season, winter (July - August), with a duration of 6 weeks each. The following parameters were evaluated: water conductivity, pH, O2 concentration, temperature, concentration of inorganic nutrients (PO4 3+, Ptotal, NH4 + , NO3 - , Ntotal), chlorophyll-a, macrophyte coverage, dry biomass, Ntotal and Ptotal of macrophytes. In both experiments, the concentrations of dissolved nutrients (PO4 3- , NH4 + e NO3 - ) in the system were reduced, but the efficiency of nutrient removal by the macrophyte was higher in summer (53% removal of P, 17% removal of N) than in the winter (23% removal of P, 4% removal of N). These results were due a higher macrophyte growth rate in summer (0.57 m2/day) than in the winter (0.35 m2/day), resulting in a higher coverage and dry biomass value. In both periods the concentration of chlorophyll-a decreased, from 70.0±36.1 μg/L to 6.9±10.0 μg/L in summer, and from 17.0±5.1 μg/L to 3.6±3.0 μg/L in winter. Thus, E. crassipes can remediate euthophication and as consequence control phytoplankton growth. The management of the macrophyte coverage at Barra do Braúna reservoir represents a potential solution to mitigate cyanobacterial blooms.
Author: Wallace Alberto da Silva de Andrade
Title: Alternative water treatment using seeds of Moringa oleifera for remotion of cyanobacteria bloom
Year: 2019 Full text (in Portuguese)
Abstract:
Moringa oleifera is a plant species native from India that has been well studied and recognized for its high nutritional value. The ability of its seeds also is known to act as a coagulant of water impurities. Currently, several studies prove the capacity of Moringa oleifera seeds to remove cyanobacteria blooms. The objective of this study was to verify the efficiency of M. oleifera seeds as an alternative water treatment for removal of cyanobacteria blooms and other microbiological contaminants. The final aim was to propose a simple and efficient methodology for the treatment of water in situations of inadequate conventional basic sanitation. Initially, an experiment was performed with jar-test equipment, using cells from the Microcystis aeruginosa species (NPLJ-4 strain). To verify the efficiency of the process, three concentrations of ground and sieved seeds of M. oleifera (10, 25 and 100 mg/L) were tested using water with different pH values (5 to 9). Water samples were harvested at various intervals (1, 6, 24 and 48 hours) to determiny turbidity (uT), concentration of chlorophyll-a and microcystins. Two other experiments were performed using raw water containing a natural cyanobacterial bloom from Funil reservoir (RJ). In both experiments, the concentration of the ground and sieved moringa seeds were 25, 50, or 100 mg/L. In the first experimente, the efficiency of the seeds was measured by the turbidity values (uT), chlorophyll-a and microcystins concentration at two intervals (6 and 24h). For the second experiment, variations of turbidity (uT), total coliforms, and Escherichia coli were analyzed. The removal of total coliforms and E. coli was not efficient. But the results showed that seeds of M. oleifera can be used as an alternative treatment to improve the quality of raw water containing natural cyanobacteria blooms. However, after the treatment with the seeds, the water needs to filter and chlorinated before consumption.
Author: Monica Hatsue Toledo Nassu
Title: Use of SODIS in the disinfection of coliforms and reduction of the concentration of microcystin-LR
Year: 2018 Full text (in Portuguese)
Abstract:
The quality of water used for human consumption is a worldwide concern, as not only pathogenic microorganisms but also toxins can be carried by this water resource. One of these toxins is microcystin, produced by several genera of cyanobacteria. In places where conventional water treatment is not available, there is a need for an alternative methodology for the treatment of this water resource, such as SODIS (Solar Water Disinfection). The SODIS methodology consists of filling transparent pet bottles with untreated water put them in the sunlight for 6 hours and let them cool for one night. After this period this water will be free of pathogens and may be consumed. To improve this technique, we can add the water to be treated, additives. The objective of the present study was to evaluate whether SODIS added or not with methylene blue (MB) and hydrogen peroxide (H2O2), could reduce the concentration of microcystin-LR and inactivate two bacterial strains, Escherichia coli (ATCC 25922) and Klebsiella pneumoniae (ATCC 13883). Our results demonstrated that none of the tested concentrations of MB, with or without the use of the chlorine tablet, exposed the SODIS methodology, were able to degrade the microcystin-LR and the blue color of the dye remained in the samples, which would make it difficult to use people. It was observed that the E. coli strain was inactivated using the SODIS methodology and with any of the H2O2 concentrations tested, whether chlorine was used or not, but the K. pneumoniae strain was only inactivated when the SODIS methodology was combined of 6 g/L of H2O2, with no use of the chlorine pellet. In relation to the concentration of microcystin-LR, in the experiments with SODIS added with H2O2, it was observed that the concentration of this toxin decreased significantly in relation to the control, being statistically equal among all tested concentrations, and close to the maximum limit allowed by the consolidation number 5/2017 Ministry of Health of Brazil. The chlorophyll-a concentration decreased significantly in all samples in relation to the control, and with SODIS use, this decrease was to below the limit of detection of the method in all samples added or not with H2O2. The results of this study are not a final solution for reducing the concentration of microcystin-LR in water for human consumption in places where conventional water treatment does not exist, but rather an initial study aiming to reduce the concentration of this cyanotoxin, using methodologies of easy handling and lower cost.
Author: Daniel Vinicius Neves de Lima
Title: Responses of Cylindrospermopis raciborskii (Cyanobacteria) strains to water conductivity effect of sodium and magnesium ions
Year: 2017. Full text (in Portuguese)
Abstract:
Semi-arid aquatic environments, such as reservoirs located at the Northeastern region of Brazil, show high water conductivity with values above 1000 μS/cm. This is associated with factors such as the mineral composition of the drainage basin and extreme climate change effects (drought, temperature and precipitation). Previous studies in these environments have shown the occurrence of cyanobacterial blooms with dominance of Cylindrospermopsis raciborskii, a potentially saxitoxin and cylindrospermopsin producer. Studies with laboratory cultivation have investigated the responses of C. raciborskii to abiotic variables such as nutrients, light and temperature, but generally include just one strain. On the other hand, it is well known that blooms consist of a “pool” of strains. Therefore to understand species’ behavior studies with more strains are required. In addition, few studies have evaluated cyanobacterial responses to water conductivity or to the stress caused by different ions (Mg2+, Ca2+, Na+). The aim of the present work was to investigate the effects of water conductivity on saxitoxin-producing and non-producing C. raciborskii strains. Four Brazilian strains were included, two saxitoxin-producers (STX+): ITEP-A1, isolated from Arcoverde reservoir (Northeast, PE), and CyRF-1, isolated from Funil reservoir (Southeast, RJ) - and two non saxitoxin-producers (STX-): CyLP, isolated from Paranoá Lake (Midwast, DF) and NPCS-1 isolated from Custódia reservoir (Northeast, PE). Strains were cultivated in ASM-1 medium and the experimental design consisted of two treatments: NaCl (10mM) and MgCl2 (5mM) addition, resulting in a conductivity of 1500 + 100 μS/cm for both conditions. Control condition was ASM-1 with no salt addition (conductivity of 450 μS/cm). Cultures were maintained at 50 μmol.photon.m2.s-1, dark: light cycle 12 hours, initial pH 8.0 and aeration during 15 days. The physiological responses of the strains were evaluated by the following parameters: growth, photosynthetic parameters, saxitoxin content, protein profiles, intracellular and extracellular carbohydrates. The NaCl treatment did not affect growth of any strain. In contrast, growth in the presence of MgCl2 varied among strains. Thus, the growth response was not related with conductivity alone but with the specific ion in solution. No differences were observed in the protein profiles and the photosynthetic parameters comparing cultures with NaCl or MgCl2 and control. Saxitoxin concentration did not change in these conditions. The production of saxitoxin did not result in advantage or disadvantage for the strains subjected to a higher conductivity. In sum, the treatments resulted in few changes in the physiology of the strains (even comparing STX+ and (STX-). On the other hand, the most evident differences were found in the individual responses of the strains maintained under the same condition. This study reveals the adaptability of C. raciborskii strains to increased conductivity, the differentiated response for each ion and the importance of considering intraspecific variability in studies of the physiology of a cyanobacteria species.
Microcystins (MCs) are cyanotoxins produced by different genera of cyanobacteria in aquatic environments. It’s a cyclic heptapeptide and toxic to many organisms. Over 90 MCs variants are described, with the –LR (MC-LR) one of the most toxic. In the Jacarepagua lagoon (RJ), cyanobacteria are abundant and reported since 1996 producing MCs. The dynamics of this molecule in the aquatic environment consists to bioaccumulate in organisms, adsorption to sediment particles and biodegradation. This study aimed to evaluate the influence of some factors such pH, atmosferic temperature and aeration, on the dynamics in the compartiments of Jacarepagua Lagoon, In addition, we evaluated the biodegradation of MC by microbial assemblages of water and sediment, characterizing them as to their bacterial composition by metagenomics. Experiments with duration of 7 days were conducted to assess both the adsorption and biodegradation activity. The parameters analyzed were not able to cause the return of preexisting toxins in the sediment to the water. Adsorption was also evaluated upon addition of a known concentration of the toxin. Acidify conditions caused an immediate reduction of the same in the water column, possibly attributed to the adsorption. Heating at 45 °C leads to reduction in the total of the toxin into the water over the 7 days, which did not occur at 25 ° C. Aeration in the water / sediment interface caused a intense decrease in the fourth day, and overall throughout the 7 days, which was not without aeration. Then, the effects evaluated may increase the adsorption capacity of the MC to sediment. However, sediment treatment with sodium azide (antimicrobial) and sterile water showed that even after handling procedures was able to degrade the toxin and thus this process has occurred simultaneously for adsorption. In biodegradation experiments by microbial assemblage (0,45 µm diameter) of water and sediment, MC was eliminated over 7 days. Microbial assemblages <0,22 µm (nanobacteria) were also evidenced and able to biodegrade the MC-LR. The characterization of water and sediment bacterial community by metagenomics showed different compositions between these compartments. Phylum Actinobacteria and Proteobacteria as were the most abundant in water, unlike interstitial water where the candidate division OP3 was the most abundant. In the raw sediment, which covers a more general community, the predominant were Chloroflexi, Proteobacteria and Firmicutes. Furthermore, it was observed of many genera described in the literature capable of biodegrading MC-LR, suggesting possible participants in the biodegradation process in the compartments of the Jacarepagua lagoon, which does not invalidate the participation of other microorganisms in general through a shared activity.
The cylindrospermopsin (CYN), a cyanotoxin, is a hydrophilic alkaloid with 415 Da. It has been described as a potent inhibitor of protein synthesis capable of causing serious damage to various organs. Studies demonstrated that the kidney was the most affected organ. Several cases of poisoning have been associated with this molecule, such as the mysterious disease Palm Island in Australia and the event in Caruaru-PE, in which patients from a hemodialysis clinic received contaminated water intravenously. Therefore, the aim of this study was to evaluate the effect of CYN in the male mice’s renal tissue. Dose-response curves with single doses intraperitoneal injection of purified CYN were performed (0, 16, 32, 64, 128, 160 µg of CYN / kg body weight) in 10 weeks male Balb/C (n = 4 animals). All animals that received doses of 128 and 160 µg of CYN/kg died within the first 72 h after toxin administration. There was no change of GFR in the 7th and the 14th days, nor in nephrin expression, a critical protein for glomerular integrity, in any of the doses. It was observed low molecular weight proteinuria and increased excretion of tubular enzymes: lactate dehydrogenase (LDH) and gamma glutamyl transferase (GGT) at doses of 16, 32 and 64 µg of CYN /kg body weight. In addition, these animals also presented respectively, increased renal interstitial space and collagen deposition, which indicates edema and fibrosis. These results suggest that CYN was able to cause a predominantly tubular damage in animals’ kidneys. Simple variance analysis (one way ANOVA) followed by Tukey's multiple comparison test were performed, using GraphPad Prism 5.0 software.
The cylindrospermopsin (CYN) - a cyanotoxin (toxin produced by cyanobacteria) - is a potent inhibitor of protein synthesis, able to cause serious damage to various organs It has the potential role of endocrine disruptor, able to inhibit the synthesis of progesterone and change the estrous cycle of females. Therefore, to analyze possible effects on males, the aim of this study was to evaluate the subchronic effect of CYN in murine male reproductive system. A subchronic assay was performed with 20 mice, with approved protocol (CEUA122/14). The animals were divided into four groups: 1, 2, 3 and control group (5 animals / group). The intraperitoneal injections at a dose of 20 μg of CYN / kg body weight occurred at the beginning of the experiment for all groups. Group 1 was euthanized and the groups 2 and 3 were re-injected 7 days after exposure to the initial dose. On the 14th day the group 2 euthanasia and reinjection of group 3 was realized. The group 3 was re-injected at 21th day and at 28th day occurred the euthanasia. Like Group 3, the control group remained until the 28th day receiving weekly injections only vehicle, ultra pure water solution (MilliQ). In the days of euthanasia (7th, 14th and 28th), blood was collected and tissues (liver, pituitary, testis and epididymis) collected and weighed. Statistical analyzes were performed with the simple analysis of variance (One way ANOVA) followed by Tukey's multiple comparison test were chosen. Our results indicate that one or two weekly low dose of CYN (groups 1 and 2) caused a significant increase in testosterone levels and there is a significant increase of sperm quantity in group 3. On the other hand, the toxin did not cause changes in the weights of the tissues as well as in the expression of pituitary (LHb and FSHb) and testicular (LHcgr, StAR and CYP11A1) genes. The changes observed have shown significant interference in the male reproductive system, although it is not known the mechanism whereby causes changes. Therefore, further studies are needed to understand the role of CYN as an endocrine disruptor.
Author: Iame Alves Guedes
Title: Diversity and Dynamics of Cyanobacteria and Microcystin production at Funil-RJ Reservoir
Year: 2013. Full text (in Portuguese)
Abstract
The Funil reservoir is located in Resende, RJ and presents frequent cyanobacterial blooms. The aim of this study was to analyze the diversity of cyanobacteria and their temporal dynamics during a bloom in the Funil reservoir, as well as the production of microcystins (MC), using different approaches. Water samples were collected from October 2011 to April 2012 and the largest values of phytoplankton biomass were detected in samples of October (5.7 mm3 .L-1 ), December (7.6 mm3 .L-1 ) and February (5.8 mm3 .L-1 ). Cyanobacteria dominated throughout the period and contributed with 88% to 99.8% of the phytoplankton biomass. The major taxa found were Microcystis spp., Cylindrospermopsis raciborskii and Dolichospermum sp. We observed the alternation of dominance between these three taxa, where Microcystis spp was dominant on October and December, C. raciborskii samples on January and Dolichospermum sp. on February samples. Phytoplankton biomass correlates positively with Microcystis biomass , NH4 + , chlorophyll and pH and negatively with NO3 - and DIN. MC was detected in all samples, including three isoforms, MC-LR, RR and YR, and concentrations ranged from 589.9 to 72.8 ng.L-1 . The proportion of Microcystis genotypes potentially producing MC was measured by qPCR and high variation was observed , ranging from 9 to 100%. Genetic diversity of cyanobacteria was evaluated by means of cpcBA sequences from each month. We sequenced 204 clones and observed 58 genotypes. Of these, 48 were genotypes of Microcystis, 7 Cylindrospermopsis 1, Pseudoanabaena and 1 Dolichospermum; 44 unique sequences were found. One Microcystis genotype was observed in all samples, 2 occurred in 5 months and the majority only in 1 or 2 months. The cpcBA marker was useful to distinguish cyanobacteria genera and to reveal intraspecific variability, but the analysis did not support species assignment in the Microcystis genera.
Author: Pedro Lima Sampaio
Title:. Allelopathic interactions between Microcystis aeruginosa and Cylindrospermopsis raciborskii isolated from Funil Reservoir, Resende (RJ).
Year: 2011. Full text (in Portuguese)
Abstract
Allelopathic interactions among phytoplankton microorganisms may play an important role in the establishment and ma intenance of algae blooms, since they can suppress the growth of potential competitors, giving competitive advantage for the allelochemical producer. Evidence suggests that allelopathy influences the shifts in dominance between Microcystis aeruginosa and Cylindrospermopsis raciborskii blooms in an eutrophic reservoir from Rio de Janeiro, Brazil (Reservatório do Funil). The aim of this study is to further investigate this possibility. For this purpose one strain of each species, isolated from the reservoir, w as used on “cross - culture” based experiments. First, a Microcystis aeruginosa strain (MiRF - 1) was grown in ASM - 1 enriched with different proportions (10%, 20% and 50%) of the exudate produced by a Cylindrospermopsis raciborskii strain (CyRF - 1). The results showed an inhibition of MiRF - 1 growth in terms of yield, only in the presence of 50% of CyRF - 1 exudate. Growth rates were not affected. Bigger colony, intense sedimentation and cellular carbohydrate accumulation were also observed in MiRF - 1 cells on this condition, but no effect on chlorophyll - a or microcystin content in MiRF - 1 occurred. Then, the effect of MiRF - 1 exudate on CyRF - 1 (50% proportionin the medium), was also tested. However, CyRF - 1 exudate did not affect MiRF - 1, regarding growth or chlorophyll - a , under the conditions tested. In addition, no evidence was found that MiRF - 1 produces an allelochemical in response to previous exposition to CyRF - 1 exudate, under the conditions tested. The results suggest that a possible allelopathic interaction betwe en Microcystis aeruginosa and Cylindrospermopsis raciborskii may influence on intense blooms and niche separation that takes place in Funil Reservoir.
Microalgae are micro-organisms promising as an alternative raw material for biodiesel production. The energy matrix is in process of change and biodegradable and renewable sources are objects of many studies nowadays. The nutritional stress is a commonly studied physiological tool to stimulate the synthesis of lipid reserves by microalgae. What is fundamental to the use of microalgae biomass for the biodiesel production. The objective of this study was to evaluate the effects of phosphorus concentration on growth and lipid synthesis by the microalgae Ankistrodesmus sp. (strain ANRF-01). For this purpose, the ASM-1 medium was manipulated relative to its initial concentration of orthophosphate. So, we obtained three different concentrations of phosphorus in the culture medium of the strain studied. Treatments appointed control, P10 and P50 started the experiment with values of 227 µM, 22.7 µM and 4.54 µM PO4 -3, respectively. The cultures were developed for 20 days at 24°C with constant aeration, light intensity of 500 μmoles photons.m-2.s- 1 , photoperiod of 12 h, initial pH 8.0 and 6 L volume of culture. Analysis were developed for cell growth, varying the concentration of chlorophyll a, carbohydrates and nutrients (nitrate and phosphate). The lipids were analyzed in relation to the percentage of crude extract and its classes of fatty acids. In the results we observed more cells and biomass for the control condition. The condition with the highest nutritional limitation (P50) showed a lower amount of biomass, but the conditions P10 and P50 showed biomass values close to the 9 th day of culture (224 and 216 mg / L, respectively). The highest values of lipids produced were detected in the P50 treatment. Among the days sampled for analysis of lipids, the 9th day of culture was the most promising for the lipid content and fatty acid profile of interest. There is need for significant amount of biomass, a high content of lipids and fatty acid profile of interest to the economical production of biodiesel from microalgae. Thus, the study indicates that the condition P50 on the 9th day (stationary phase) would be more appropriate as an alternative source, in a future production of biodiesel on a large scale. Further studies should be conducted to give a better basis on the physiological conditions of cultivation of these organisms for the biotechnology purpose.
Author: Luana Jotha Mattos
Title: Sublethal effects of microcystin (cyanotoxin) on biochemical, physiological and stereological aspects in murine models.
Year: 2011. Full text (in Portuguese)
Abstract
Microcystins (MCYST) are the cyanotoxins with more reports of human poisoning, but the sublethal dose effects of these toxins are poorly studied. Therefore, the aim of this study was to contribute to knowledge about the effects of intoxication by MCYST-LR (sublethal dose) in mammals, investigating biochemical and physiological changes, processes of accumulation and detoxification, as well as investigate the functional and tissue damage in the exposed animals. We used mice and rats as experimental models. The mice were exposed to MCYST-LR i.p. with single sublethal dose of 45 mg MCYST-LR / kg bw. Their organs were collected at 2, 8, 24, 48 or 96 h. The rats were injected (IP) with single sublethal dose of 55 mg MCYST-LR / kg BW and their organs were collected after 24 h. In all experiments, control animals received saline solution. In mice, we could detect oxidative damage in the liver, particularly in 8 h after exposure, justified by the increased activity of SOD (superoxide dismutase), decreased activity of CAT (catalase) and GPx (glutathione peroxidase) and increased lipid peroxidation. Inhibition of protein phosphatase (PP) occurred during the experimental period. Analysis of nitric oxide and myeloperoxidase were not considered as good biomarkers of inflammation. The activity of GST (glutathione S-transferase) increased at early sample days, indicating detoxification. The biomarkers of liver injury were detected in high concentrations in the serum of animals exposed during the sample period. Stereological analysis indicated an initial moment of intoxication, justified by the increase of steatosis, and a later moment of intoxication, justified by the increase of inflammation and hepatocytes binucleation. Necrosis and increase in blood vessel diameter were observed during the experimental period. The volume and number of hepatocytes per unit area also decreased, but it was possible to verify the recovery of these parameters over time of exposure. The MCYST has accumulated in hepatocytes and can be detected until the last analysis time. In the liver of rats, the same histopathological pattern was observed in 24 h. A possible oxidative damage was found in liver and kidney justified by the change of CAT activity and the relationship GSH / GSSG. The kidney index in rat was increased at 24 h, indicating a possible edema. It was observed that the rats intoxicated decreased food intake and feces production, and increased of drink ingestion and elimination of urine. Similarly, the glomerular filtration rate increases in animals exposed to toxin. There was a greater elimination of MCYST in the free form in the urine in 24 h, even though only 14% of the administered dose had been eliminated. However, the fractional excretion of the same was 137.9 ± 9.1%, indicating secretion process. The results of this study confirm the presence of MCYST-LR in mice exposed to a single dose of MCYST. In addition, there were metabolic and functional alterations in the kidneys of rats exposed. These results confirm the necessity for more studies despite of sublethal exposures to MCYST.
Author: Laís Bessa dos Santos
Title: Effects of phosphorous concentration on growth and lipid synthesis in Ankistrodesmus sp.
Year: 2011. Full text (in Portuguese)
Abstract:
Microalgae are micro-organisms promising as an alternative raw material for biodiesel production. The energy matrix is in process of change and biodegradable and renewable sources are objects of many studies nowadays. The nutritional stress is a commonly studied physiological tool to stimulate the synthesis of lipid reserves by microalgae. What is fundamental to the use of microalgae biomass for the biodiesel production. The objective of this study was to evaluate the effects of phosphorus concentration on growth and lipid synthesis by the microalgae Ankistrodesmus sp. (strain ANRF-01). For this purpose, the ASM-1 medium was manipulated relative to its initial concentration of orthophosphate. So, we obtained three different concentrations of phosphorus in the culture medium of the strain studied. Treatments appointed control, P10 and P50 started the experiment with values of 227 μM, 22.7 μM and 4.54 μM PO4 -3, respectively. The cultures were developed for 20 days at 24°C with constant aeration, light intensity of 500 μmoles photons.m-2.s- 1, photoperiod of 12 h, initial pH 8.0 and 6 L volume of culture. Analysis were developed for cell growth, varying the concentration of chlorophyll a, carbohydrates and nutrients (nitrate and phosphate). The lipids were analyzed in relation to the percentage of crude extract and its classes of fatty acids. In the results we observed more cells and biomass for the control condition. The condition with the highest nutritional limitation (P50) showed a lower amount of biomass, but the conditions P10 and P50 showed biomass values close to the 9 th day of culture (224 and 216 mg / L, respectively). The highest values of lipids produced were detected in the P50 treatment. Among the days sampled for analysis of lipids, the 9th day of culture was the most promising for the lipid content and fatty acid profile of interest. There is need for significant amount of biomass, a high content of lipids and fatty acid profile of interest to the economical production of biodiesel from microalgae. Thus, the study indicates that the condition P50 on the 9th day (stationary phase) would be more appropriate as an alternative source, in a future production of biodiesel on a large scale. Further studies should be conducted to give a better basis on the physiological conditions of cultivation of these organisms for the biotechnology purpose. Key words: microalgae, lipids, phosphorus concentration and biodiesel.
Author: Carolina Tolomini Miranda.
Title: Effects of light intensity in growth and lipid production by Akistrodesmus sp. (Chlorophyceae) for biodiesel production.
Year: 2011. Full text (in Portuguese)
Abstract:
The variation in light intensity can promote changes in morphology and photosynthetic characteristics of microalgae. However, is still unclear if high light intensities could have a role in lipids production. The aim of this study was to investigate the variation of lipids accumulation during growth of a strain of Ankistrodesmus sp. (ANRF-01) under different light intensities: 900, 1400 and 2000 μmols photons.m-2.s-1. Cultures were grown during 20 days in bath culture conditions, ASM-1 medium, at 24±1ºC, pH 8 and light\dark cycles of 12 hours. Sampling was done every two days for analysis of cell number, chlorophyll-a and carbohydrates concentration. Lipid production was determined at: 4th (exponential phase), 8th (early stationary phase), 12th and 19th (stationary phase) days. The maximum yield of increase (defined as maximum cells number divided of the inoculum cells) was 0,62 in 900 μmols photons.m-2.s-1. In 1400 and 2000 μmols photons.m-2.s-1 these rates were 0,59 and 0,58 respectively. Biomass production was highest at the lowest light intensity (894 ± 53mg.L-1). These values were 841 ± 40.3mg.L-1 for 1400 μmols photons.m-2.s-1 and 771 ± 3.2mg.L-1 for 2000 μmols photons.m-2.s-1. In contrast the highest percentages of lipid extracts were obtained in 2000 μmols photons.m-2.s-1, reaching 46 ± 7.3% of biomass, while in 900 μmols photons.m-2.s-1 it reached 40 ± 4% and 33 ± 7,8% in 1400 μmols photons.m-2.s-1. The highest yield of fatty acid esters (68 ± 20.5%) were obtained in 1400 μmols while in 900 and 2000 μmols photons.m-2.s-1 were 37±4.4 and 33±5.7% respectively. Gas chromatography analysis showed that ester profile is adequate to biodiesel production. Data showed that light intensities usually found in surface water column are high enough to allow a good biomass and synthesis of fatty acid esters adequate to biodiesel production.
Author: Maria Elisângela Venâncio dos Santos.
Title: Effect of exudate in physiological autoregulation of Microcystis sp.
Year: 2009. Full text (in Portuguese)
Abstract:
Effect of exudates on the physiological self Microcystis sp. Quorum sensing is a process of cell communication that plays a central role in the physiology of microorganisms. Many bacteria use this system and coordinate their activities in a manner similar to multicellular organisms. This study aimed to investigate whether cyanobacteria use signaling molecules of quorum sensing to regulate their physiology, and the entry of the population in stationary phase of growth, as already observed in other bacteria. For this study, a strain of Microcystis sp. (Milj-48) was grown in media previously conditioned by itself (exudatos), obtained from different stages of growth. To evaluate the effect of higher concentrations of exudato, was also an experiment with the addition of exudatos focused on 100 and 50x for crops. The effects produced by exudatos was verified by assessment: cell growth, by counting the cells, the variation of the concentration of chlorophyll a and varying the concentrations of microcystins, using the method of enzyme immunoassay (ELISA) for detection of total microcystins and High Performance Liquid Chromatography (HPLC) for detection of intracellular microcystin. It demonstrated an inhibition of cell growth with all exudatos tested on the control. The most pronounced effect was obtained with exudato the 6th day of growth (phase exponencial). There was no effect by increasing the concentration of this exudato. The cultures showed no inhibition in the production of chlorophyll a. The addition of the exudato cultivation resulted in cell aggregation and chlorosis, phenomena typical of cultures in stationary phase or in limitation of nutrients. A positive correlation between production of toxin and inhibition of growth is apparent. It is suggested that the strain of Microcystis sp. (Milj-48) promotes the release of a yet unknown signal that interferes with their own cell division, we raise the possibility of involvement of this signaling microcystin.
Author: Priscila Rodrigues Koschek
Title:.Interaction between Monoraphidium and Microcystis using medium with released organic matter and mixed cultures
Year: 2008. Full text (in Portuguese)
Abstract
Interspecific communication has an important role in phytoplankton growth and this may be true also for toxin production by cyanobacteria. This group of microorganisms is frequently found as dominant due to eutrophication processes in aquatic environments. Data from Funil Reservoir (Resende-RJ) have demonstrated that at least 90% of phytoplanktonic biomass can be composed of cyanobacteria. Microcystis is one of the main dominant genera. The present study was designed to investigate the interaction between one strain of the chlorophyte Monoraphidium, isolated from that reservoir, and four strains of Microcystis. Cultivation using medium with released organic matter (exudate) from monospecifc cultures and also from mixed cultures, in different proportions, was performed. Mixed cultures included the chlorophyte and one toxic strain of Microcystis (MIRF-01) isolated from the same environment or another nontoxic strain of Microcystis (MICD-01) isolated from a different environment. Physiological conditions of strains were followed by cell counting and by analysis of chlorophyll a. The variation in microcystins production was evaluated by HPLC technique. The results showed that medium with exudates from monospecific cultures of Microcystis did not inhibit growth of Monoraphidium with aeration, except for the 10% concentration exudates, and that occurred in one of the two strains of Microcystis tested. However, in cultivation without aeration the chlorophyte had its growth stimulated by exudate (50%) from one in three Microcystis strains tested. In addition, this stimulatory effect also involved reduction in chlorophyll a. The addition of exudates from Monoraphidium induced growth of one in two Microcystis strains tested, without changes in chlorophyll a. In conditions of mixed cultivation, Monoraphidium as well as Microcystis showed reduced growth as compared to monospecific cultures and the chlorophyte showed increased growth with Microcystis MIRF-01 strain as compared to MICD-01 strain. In the presence of 100% exudate from mixed cultivation inoculated with 70% Monoraphidium and 30% Microcystis the growth of Monoraphidium was inhibited with reduction of chlorophyll a, characterizing chlorosis at the end of cultivation (6 days). For Microcystis, the addition of this same exudate (50%) tended to inhibit cellular growth without affecting chlorophyll a. Microcystins concentration did not change in mixed cultures with Monoraphidium, however it increased using medium with exudates from Monoraphidium monospecific cultures or from mixed cultures in which Monoraphidium predominated. Although the function of microcystins in this interaction is still not clear, this study contributes to enforce the importance of infochemichals for this cyanotoxin production. Therefore, data show that organic matter released by these species varies in relation to growth conditions. Besides, it may have a controlling role in their metabolism at inter-specific and possibly intra-specific level.
Author: Rodrigo de Cássio da Silva
Title:.Bioaccumulation and depuration of the cyanobacterial toxin cylindrospermopsin and its effects on growth in the juvenile tilapia (Oreochromis niloticus)
Year: 2008. Full text (in Portuguese)
Abstract
Cylindrospermopsin (CYN) is a toxic alkaloid produced by several genera of cyanobacteria and is implicated in human and wildlife poisoning incidents. The principal mode of action for CYN is protein synthesis inhibition, but the sublethal effects of this toxin in aquatic organisms, including changes to development and behavior, have been poorly studied. Little information is available regarding bioaccumulation of CYN in these organisms. This study examined its bioaccumulation, depuration and effects on growth in the juvenile tilapia (Oreochromis niloticus) exposed to freeze-thawed whole cell extracts and whole cells of toxic (Cylindrospermopsis raciborskii). Four experiments under laboratory conditions with these fish were done. In the first one the animals were fed for 15 days with fish food plus cells extracts (0.5 g food.day-1. fish-1) containing 0.31 μg CYN .g-1 food. Every three days for a fifteen day period, fish were collected. In the second trial, fish were fed during 30 days and the fish food (0.5 g food.day-1. fish-1) contained 0.35 μg CYN .g-1 food. After 15 days, fish were collected each 3 days. In the third trial fish received (0.5 g food.day-1. fish-1), during 12 days, fish food with cells extract at a higher toxin concentration (0.8 μg CYN .g-1 food). In the following 10 days, the animals were fed (0.5 g food.day-1. fish-1) without toxic cells extracts. The fourth trial, fish received (0.5 g food.day-1. fish-1), during 15 days, food plus whole cells of C. raciborskii (5.4 μg CYN .g-1 food). Fish weight and length were measured, its samples of muscle tissue and viscera, in all trials, were extracted with water and 100% methanol, filtered, evaporated and resuspendend in water and analyzed by ELISA imunoassays. It was verified that CYN exposed animals had their biomass and sizes reduced in relation to non-exposed fish in the trial 1, but not in the others trials. In the first trial the higher concentration of CYN in muscle tissue was observed on third day, tending to reduce on day with oscillations throughout the time. Viscera showed the maximum concentration on third day and presented the same concentration oscillation pattern throughout the time. In the second trial, maximum concentration was found in the first sampling day on muscle tissues (19th day – 0.15 ng CYN.g-1) and viscera (7.83 ng CYN.g-1). In the depuration experiment (3rd trial) the concentration found in muscle tissue and viscera, in depuration period, were almost the same found on first sampling day. In the last trial (4th trial), highest concentrations of CYN in muscle tissue was found on 12nd day (2.3 ng CYN. g-1). But in viscera, the higher concentration was found on 15th day (21.1 ng CYN. g-1). Studies of CYN distribution in animal tissues showed that the majority of this toxin is observed in viscera, resulting from the consumption of toxin-containing food. In this study we have noted a bi-phasic pattern of CYN throughout the time and it could be attributed to the detoxification process. These results do not corroborate other works since fish accumulated CYN when nourished with cellular extracts more than toxic cells. This is the first investigation of CYN bioaccumulation in tilapia and indicates the importance of continuous monitoring of this compound in organisms for human and animal consumptions.
Author: Maria Isabel de Almeida Rocha
Title:.Seasonal variation of phytoplankton and zooplankton and the concentration of microcystins in these two trophic levels in the Reservoir Funil (Resende-RJ)
Year: 2007. Full text (in Portuguese)
Abstract
The increase of nutrients input in lakes and reservoirs linked to different environmental factors accelerates the eutrophication process and establishes ideal conditions for cyanobacterial blooms. Funil Reservoir located between São Paulo and Rio de Janeiro States, formed by Paraíba do Sul River, was built in an industrial and agricultural region, with propitious characteristics for development of potentially toxic cyanobacterial bloom. The aim of this work was to evaluate the occurrence and transference of microcystins in the phytoplanktonic and zooplanktonic community of this reservoir, considering the composition and variation of both. Moreover, physical and chemical analyses of water were done monthly, throughout two consecutive years. Phytoplankton and zooplankton communities were previously separated, freeze-dried and, for the microcystins extraction, solubilized in acidified methanol. The extract was cleaned-up and analyzed by ELISA. Physical and chemical analyses demonstrated that the temperature of water varied approximately from 20 – 30ºC, pH had neutral-alkaline characteristics, the average of the dissolved oxygen concentration was 6.82 mg l-1 and the conductivity was 96.9 μS cm-1. These values, together with raised values of nutrients that arrive to Funil Reservoir, give adequate conditions to formation of a cyanobacterial bloom, causing reduction of the transparency of the water. The phytoplanktonic community was dominated by cyanobacteria, which reached more than 90% of the density in almost every month of study. The predominant genus was Microcystis, as in colonial or unicellular forms. Zooplankton varied monthly in density and composition of species. The copepods group had the highest densities and the rotifers group contributed with the highest variety of species. The population of cladocerans was reduced, with the dominance of the smallest one. These alterations in the zooplanktonic community can be associated to the constant presence of cyanobacterial bloom, which favors species already adapted to survive in these conditions. During all period microcystins were found in phytoplankton samples, reaching the maximum of 3.789 μg g-1, in December, 2004. These microcystins have been transferred to zooplankton communities, with values two orders of magnitude lower than phytoplankton approximately, with maximum of 63.15 μg g-1, in October, 2004. In accordance to these microcystins concentration results, it was observed three distinct phases: the first one with low microcystins concentration and low phytoplankton density; the second one with variation in microcystins concentration and in Microcystis density, and the third one with microcystins concentrations close to the first phase, but with an increase in phytoplanktonic population density. This is indicating, probably, the occurrence of three distinct populations. Considering these conclusions, the zooplanktonic community of the Funil Reservoir cannot be considered as a good vector of microcystins for higher trophic levels.
Author: Andreia Maria da Anunciação Gomes.
Title: Impact of intensive fish farming activity and the addition of inorganic nutrients (N and P) in water quality in the reservoir of Ribeirão das Lages-RJ.
Year: 2005. Full text (in Portuguese)
Abstract
This study aims to determine the influence of inorganic nutrient addition (N and P) and fish culture on physical, chemical and biologic features of the water of Ribeirão das Lajes Reservoir, a oligo-mesotrophic system. This research was conducted through in situ experiments performed during february 2004. For this purpose we used 9 polyethylene bags (2900 liters), placed in the reservoir at the same time. The enclosures were divided in three treatments: one of them containing fishes (10 tilapia per enclosure); the second of them with nutrient addition (nitrate, ammonium and phosphorus addition with amounts of 70; 1,3 and 1,2 μg/L/day, respectively) and the third of them containing no fishes and no nutrient addition (control group). Water samples were collected from each enclosure and from the reservoir in alternated days until the 15th day and the last ones were collected 25 days after the beginning of the experiments. The water temperature, pH, conductivity and turbidity were measured in situ with specific electrodes. Secchi depth also was determined. Chlorophyll a and nutrient concentrations were measured spectrophotometrically. Algal numbers were determined with an inverted microscope from lugol-fixed samples. During experimental periods, oligotrophic conditions characterized the reservoir, like: low concentration of nitrogen and phosphorus, high secchi depth, low turbidity and low phytoplankton density. Phytoplankton community was dominated by cyanobacteria in Ribeirão das Lajes reservoir. The control group remained these features. Nutrient addition caused an increase of nitrate concentration, but the P addition was not sufficient to increase SRP concentration. Although Chlorophyta had replaced Cyanobacteria, the phytoplankton density was not increased in these enclosures. However, changes related to the trophic conditions had taken place in the enclosures containing fishes. The tilapia presence increased water temperature, pH, conductivity, turbidity values and secchi depth values. The chlorophyll increased from 1,5μg/L, at the beginning of the experiment, to 542,28 μg/L at the end of experiment. The growth of algae leads to decreases of the transparency level. Phytoplankton community structure was changed in enclosures containing fishes. Cyanobacteria were excluded from these enclosures and Chlorophyta were dominant at the end of experiment. The present study results suggest that nutrient addition was not sufficient to increase phytoplankton density and the major limiting factors for the phytoplankton growth could be phosphorus deficiency. However, the high nitrogen concentrations observed in all treatment enclosures were sufficient to change the phytoplankton community structure, leading chlorophyta increase. Chorophyta dominance in spite of cyanobacteria has been observed in hypereutrophic system.
Author: Ronaldo Leal Carneiro
Title: Optimization of cultivation of Microcystis aeruginosa and Cylindrospermopsis raciborkii (Cyanobacteria) for production of microcystin-LR and cylindrospermopsin standards.
Year: 2005. Full text (in Portuguese)
Abstract
Hepatotoxinas microcystin-LR and cylindrospermopsin produced by cyanobacteria have already been associated with cases of animal and human intoxication. The Brazilian Ministry of Health adopts as the acceptable maximum level 1 μg/L of microcystins in water for human consumption and recommends 15 μg/L of cylindrospermopsin. Additionally, drinking water for public provisioning must been monitored for the presence of cianotoxinas. This study aimed to collaborate with the establishment of optimized conditions for production of these two cianotoxinas to allow the possibility of the national production of microcystin-LR and cylindrospermopsin standards, which is crucial for water quality control process. We studied the variation on microcystin-LR synthesis, by a strain of Microcystis aeruginosa (NPJB-1) and in the cylindrospermopsin synthesis, by a strain of Cylindrospermopsis raciborskii (CYP 011K) in batch cultures submitted to different irradiance levels. Growth rate and physiological conditions of each strain were determined by daily cell counting, chlorophyll a concentration (extraction with methanol 100%), intracellular and extracellular carbohydrates concentration (using the phenol sulfuric acid method). The production of these two toxins was quantified in the exponential and stationary growth phases by specific HPLC techniques using UV detection and photodiiodearray detector. The best conditions for growth and microcystin-LR production in 106 cells of strain NPJB-1 (M.aeruginosa) were found in the cultures maintained under 100 μmoles fótons.m-2.s-1. The age of culture influenced in the cellular microcystin-LR concentration. The content was higher in the exponential phase and it decreased in the stationary phase. There was a direct relationship between chlorophyll a synthesis and microcystin-LR production by this strain. According to these results, the most favorable culture conditions for production of microcystin-LR standards by NPJB-1 strain are 100 μmoles fótons.m-2.s-1 for five days of cultivation. The highest growth rate of CYP 011K (C. raciborskii) strain was found in the cultures maintained under of 100 μmoles fótons.m-2.s-1. A viii .s-in the middle of exponential growth phase as well as in the end of culture period. According to these results, the most favorable culture conditions for production of cylindrospermopsin standards are 40 μmoles fótons.m-.s-for twelve days of cultivation. decrease of the length of C. raciborskii filaments was observed at the end of exponential growth phase for the cultures submitted to the highest light treatments (60 and 100 μmoles fótons.m-2.s-1). The highest cylindrospermopsin production in 106 cells of C. Raciborskii was observed in the cultures submitted to 40 μmoles fótons.m-21 21
Author: Raquel Moraes Soares
Title:.Accumulation, depuration and effects of mycrocystins (cyanobacteria hepatotoxins) in Tilapia rendalli (Cichlidae)
Year: 1999. Full text (in Portuguese)
Abstract
The occurrence of blooms of Microcystis aeruginosa in several water bodies has been frequently reported. This cyanobacteria species is able to produce microcystins hepatotoxic heptapeptide), which can cause serious damage on the aquatic biota and mammalian. The effects of the toxic cyanobacteria M aeruginosa in fish are relevant due to the ecological, economic and social importance of these organisms usually consumed as human food. This work studied the process of accumulation and depuration of microcystins in Tilapia rendalli and the effects of these toxins on the growth of this fish. Three experiments with juveniles maintained in lab conditions were done. In the first one each fish received, during 15 days, fish food plus toxic cells of M aeruginosa (20.4µg of microcystins per day). In the following 15 days, the organisms were feed without toxic cells. Four fishes were harvested every 3 days and feces were collected for microcystins analyses. In the second experiment, fishes were feed, only with toxic cells of M aeruginosa (I 4,6J.1g of microcystins/day/fish) during 28 days. In the third experiment, fishes were feed with fish food plus toxic cells previously lised (29,2J.1g of microcystins/day/fish), during 6 weeks. In these two last experiments, fishes were harvested every 7 days, and the biometric parameters were annotated. The microcystin concentration in the liver, muscle and fasces were determined by ELISA immunoassays. In the first experiment the highest concentration of microcystins in the liver tissue was observed on the 6th.day (0,6J.1g.g-1), while in the muscle it occurred during depuration period (24th. day 0,05J.1g.g-1). It suggested a possible transference of microcystins from the liver to the muscle. Almost 50% of the toxins were released through the fasces during the depuration period. In the second experiment, the nourishment only with toxic cells could have caused the accumulation of higher microcystin concentrations •than in the other experiments. In the third experiment, the toxins were released by cellular lise and the uptake of these toxins seemed to be different. In the liver, the highest concentration was observed on the last day (1,7J.1g.g-1). During this experiment a decrease in body weight was verified in the intoxicated individuals. This study showed us that Tilapia rendalli when consuming M aeruginosa cells and microcystins in concentrations usually found during blooms occurrence can accumulate these toxins above the tolerable limit for human consumption. This fact can be a serious risk for aquatic ecosystem and public health.
Author: Simone Maciel da Costa Gouvea
Title: Antimicrobial activity of compounds isolated from strains of cyanobacteria family Oscillatoriaceae.
Year: 1998. Full text (in Portuguese)
Abstract
Biological activity of secondary metabolites produced by cyanobacteria have been studied due to their importance in biotechnology and/or pharmacology. These metabolites have a wide spectrum of activity: while: allelopathic, algicidal; antibacterial, antifungal, antineoplastic, cytotoxic, antiviral, enzyme inhibitors and inflammatory. Biotechnological potential of polar extracts of some strains of cyanobacteria (Oscillatoriales) of Brazilian water bodies were evaluated through the following biological tests: diffusion tests, platelet aggregation, antiviral and cytotoxic activity. In diffusion tests, the better results were with MeOH 80% (N) extracts of the strains NPCD-2 (Limnothrix sp), NPDF-2 (Leptolyngbya sp) and NPBT-l (Tychonema sp) strains against Candida albicans. However the extract lost activity when the strains were subcultured for long periods, probably due to a decreased production of the bioactive metabolite. In the platelet aggregation tests, the extracts (N) and (I) of NPFX -1 - Leptolyngbya sp (MeOH 20%), NPFE-l - Tychonema sp (MeOH 20%) and NPCD-2- Limnothrix sp (H20) strains, were able to induce a natural platelet aggregation in the absence of the agonists ADP and P AF. This result suggests indirectly a potential inflammatory activity of these extracts. Nevertheless antiviral tests showed no activity against HSV -1 (Herpes simplex virus - type 1). Cytotoxic tests showed that the aqueous extracts of NPRG-l, NPDF-2 and NPRE-3 strains, both Leptolyngbya sp (Pseudoanabaenaceae) genera, can provide important information to developed new anti-cancer drugs. These results confirm the potential of extracts of cyanobacteria from the order Oscillatoriales as a promising source of new bioactive metabolites and could provide subsidy to future investigations chemical and biotechnological.
Author: Ana Cláudia Pimentel de Oliveira.
Title: Ecophysiological studies of Synechocystis aquatilis f. aquatilis (Cyanophyceae)and microcystin production in unialgal and mixed cultures
Year: 1997. Full text (in Portuguese)
Abstract
Biological activity of secondary metabolites produced by cyanobacteria have been studied due to their importance in biotechnology and/or pharmacology. These metabolites have a wide spectrum of activity: while: allelopathic, algicidal; antibacterial, antifungal, antineoplastic, cytotoxic, antiviral, enzyme inhibitors and inflammatory. Biotechnological potential of polar extracts of some strains of cyanobacteria (Oscillatoriales) of Brazilian water bodies were evaluated through the following biological tests: diffusion tests, platelet aggregation, antiviral and cytotoxic activity. In diffusion tests, the better results were with MeOH 80% (N) extracts of the strains NPCD-2 (Limnothrix sp), NPDF-2 (Leptolyngbya sp) and NPBT-l (Tychonema sp) strains against Candida albicans. However the extract lost activity when the strains were subcultured for long periods, probably due to a decreased production of the bioactive metabolite. In the platelet aggregation tests, the extracts (N) and (I) of NPFX -1 - Leptolyngbya sp (MeOH 20%), NPFE-l - Tychonema sp (MeOH 20%) and NPCD-2- Limnothrix sp (H20) strains, were able to induce a natural platelet aggregation in the absence of the agonists ADP and P AF. This result suggests indirectly a potential inflammatory activity of these extracts. Nevertheless antiviral tests showed no activity against HSV -1 (Herpes simplex virus - type 1). Cytotoxic tests showed that the aqueous extracts of NPRG-l, NPDF-2 and NPRE-3 strains, both Leptolyngbya sp (Pseudoanabaenaceae) genera, can provide important information to developed new anti-cancer drugs. These results confirm the potential of extracts of cyanobacteria from the order Oscillatoriales as a promising source of new bioactive metabolites and could provide subsidy to future investigations chemical and biotechnological. The occurrence of toxic 'blooms' of Cyanophyceae in eutrophic lakes, rivers and reservoirs has been frequent. Cyanophyceae toxins may cause poisoning of mammals, birds, fishes and several effects in public health. Hepatotoxins are the most common toxins produced by cyanophyceae and have been receiving more attention due to their biological activity and wide distribution. Nevertheless, the controlling mechanisms of cyanotoxins production has not been fully understood yet, although there are already some evidences of dependency on nutritional factors for the stimulation of growth and synthesis of these hepatotoxins. Previous studies with the strain NPBS-2 of Synechocystis aquatilis f aquatilis have shown that this strain presented a high degree of hepatotoxicity when grown in mixed cultures with the Chlorophyceae Monorraphydium convolutum. Due to this fact, experiments with unialgal and mixed cultures were carried out with both species with the aim of understanding which are the likely factors that would be stimulating the production of microcystins by Synechocystis aquatilis f. aquatilis. The cultures of both species were carried out in ASM-I and Z-8 standard media and in modified 'Special' and 'Reconstituted' media which have the nutritional composition modified from ASM-I and Z-8 media respectively. During the experiments, we followed the physiological state of the cultures by cell counting and by analysis of chlorophyll-a, intra and extracellular carbohydrates and protein. The variation in microcystin production was evaluated by mouse bioassay, immunoassay and High Performance Liquid Chromatography (HPLC). The cultures carried out in Z-8 medium, which is three times more concentrated in nitrate than ASM-l medium, showed a good cellular growth. However, in this culture medium, cells grown in unialgal and mixed conditions have shown no toxicity. Nevertheless, the mixed cultures carried out in ASM-I medium showed high toxicity. The chemical composition of the culture media significantly affected the growth of both species in unialgal and mixed cultures. Microcystins production also varied under these different culture conditions. The chromatograms obtained by HPLC showed that S. aquatilis f aquatilis grown in different proportions of M. COI1VO/utlln7 presented a variation in both quality and quantity of microcystins production. Therefore, one must consider that besides the physical and chemical factors already determined in other studies, the cellular growth of cyanophyceae and microcystins production by these organisms may be affected by biological factors related to interspecific interactions.
Author: Silvia Mattos Nascimento.
Title: An ecophysiological study of Synechocystis aquatilis f. salina: Ratio N / P and its relation to the growth and photosynthetic activity
Year: 1996. Full text (in Portuguese)
Abstract
Over a 3-year period (October 1990 to August 1993) cyanobacteria represented 60% of the density and 52% of the biomass of the phytoplankton community of Lagoa da Barra (Maricá-RJ, Brazil). It was also seen that the cyanobacterial biomass was negatively correlated with nitrate+nitrite concentrations and DINIP04, PONIPOP, N03ITP e TN/TP ratios. Synechocystis aquatilis f salina was the predominant species, representing 57% of the cyanobacterial density during the entire period of the study Based on this data, the species was cultivated under laboratory conditions in ASM-I medium and ASM-I 10 times diluted for ail nutrients (ASM-1I10) with the following NIP ratios, 3:1, 10:1 and 30:1. Growth rate, chlorophyll a, protein and carbohydrate intracellular concentrations, carbohydrate excretion and the photosynthetic activity were investigated. It was ascertained that the species is well adapted to a condition of nitrogen limitation, compared to the Redfield ratio. High chlorophyll a and protein concentrations as well as the highest photosynthetic rate were observed on cultures of ASM-I medium with low NIP ratio (3: I). Furthermore, the cultures in medium with the highest NIP ratio (30: I) showed that the species is not adapted to this condition. This study is a contribution to the knowledge of the strategies utilized by cyanobacteria, which gives them a competitive advantage under nitrogen limitation.
Author: Renato José Reis Molica.
Title:Effect of light intensity on growth and microcystin production in two strains of Microcystis aeruginosa Kutz Emend. Elenkin (Cyanophyceae)
Year: 1996. Full text (in Portuguese)
Abstract
Accelerating eutrophication of waterbodies has resulted in an exaggerated growth of blue-green algae or cyanobacteria all over the world, a phenomenon named bloom. Moreover, some of the species that form blooms are toxins-producers and are responsible for several cases of animal death and human illness. Hepatotoxins are the toxins more related to these cases and, within this group, microcystin remains the main representative. Among the several species causing toxic water blooms, Microcystis aeruginosa is the most common. During blooms, cells are submitted to high light intensities. In order to examine how microcystin production can be altered by this condition, two strains of M aeruginosa (NPLJ-4 and NPCD-1, toxic and non-toxic, respectively) were cultivated under 40µmol photons .m•2.s-l , 180µmol photons.m•2.s-l and subjected to a change in irradiance from 40 to 180µmol photons. m•2.s-l, when the cultures reached the period between the exponential and the stationary growth phases and the late exponential phase. The physiological state of the cells was accompanied by cell counting and by the analysis of some physiological parameters: chlorophyll a, carbohydrate, protein and biomass production. Also, photosynthetic light-response curves of the NPLJ-4 strain were developed when the irradiance was altered at the late exponential growth phase. Changes in light intensity did not induce toxin production by the non-toxic strain (NPCD-1). The growth phases and the different irradiance regimes influenced the microcystin synthesis by the toxic strain (NPLJ-4). The intracellular level of microcystin was highest during the period between the exponential and the stationary phases, and decreased during the stationary phase. At light intensity of 40 µmoll photons. m•2.s-l the cells contained the highest amounts of toxin and the change in irradiance to 180µmol photons.m•2.s-l caused a decrease in microcystin levels. The change in light intensity resulted in an increase in cell division. The chlorophyll, carbohydrate and protein levels tended to equalize those of the cultures that had grown at 180µmol photons.m•2.s-l. The photosynthetic light-response curves showed that cells submitted to a change in irradiance were photoinhibited and the highest values of light saturated photosynthetic rate were observed in the cultures cultivated at 180µmol photons.m•2.s-l. Our experiments did not a1low to conc1ude how light controls microcystin synthesis. However, hypothesis could be speculated: at high light intensities, most of the energy could be detected to enhance the efficiency of CO2 absorption, consequent1y, the synthesis of the enzymes responsible for microcystin synthesis diminished. The other one could be that the decrease of microcystin levels, associated with the increase of light intensity, would be a consequence of the rise in growth rate.
Author: Débora Guerreiro Aguiar.
Title: Effect of different nutrient concentration on growth and toxin production by Microcystis aeruginosa Kütz Emend. Elenkin (Cyanophyceae) in culture conditions.
Year: 1995. Full text (in Portuguese)
Abstract
The increase of eutrophication of Brazilian natural freshwater supplies and reservoirs has led to Cyanophyta species becoming dominant. The most common Cyanophyta genera producing "blooms" on the surface of eutrophic Brazilian water bodies is Microcystis and the hepatotoxin production by strains of this genera has been showed in many areas of the world In order to understand the relationship between nutrient concentration and hepatotoxin production, a hepatotoxic strain (NPJB- l) of Microcystis aeruginosa, isolated from Lagoa das Garças (São Paulo city - Brazil) was cultivated in ASM-l medium using different concentrations of the nutrients (N/20, N/10, N/5, ASM-l (C), 2N, 2NP, 5N, 5NP, ASM-l/2, 2ASM-l and 5ASM-l) at 22uE om-2 o S-1 of light with a photoperiod of 12 h and 2l±2°C of temperature. The culture growth was monitored by cell count, growth rates, biomass production and cell productivity, Cells growing on the exponential lyophilized phase were harvested, for mouse bioassays and concentrated and toxin extraction o isolation, purification and quantification of the toxins were carried out by reverse phase HPLC o The growth rate was reduced in the conditions 2N, 5N and ASM-l/2 and it was increased under 2ASM-l and 5ASM- l conditions. The cell biomass and cell productivity were increased in the conditions 2ASM- l and 5ASM- lo Cell productivity was reduced under N/20, N/10, N/5 and ASM- l/2 conditions. This Microcystis s aeruginosa strain (NPJB-l) produces two different hepatotoxic heptapeptide microcystins: LR and LF, however the decrease of hepatotoxicity under the conditions ASM-l/2 and 2ASM-l was mainly related with the decrease of microcystin- LR production. The best culture condition for the growth of this strain was not coincident with the condition where was observed the maximum hepatotoxins production. Therefore the increase of nutrient concentration in aquatic environment should not be directly related with the hepatotoxin production by the Microcystis strains.
Author: Cláudio Roberto Ribeiro Bobeda.
Title: Isolation, purification and characterization of microcystins (heptapeptides toxic) found in cyanobacterial blooms occurred in the reservoir Funil (Resende, RJ).
Year: 1993. Full text (in Portuguese)
Abstract
Several cyanobacteria species (blue-green algae) that I usually present mass development in fresh and brackish waters have been described as responsible for animal and human intoxication in many countries. Although there are no confirmed poisoning cases of animals or humans by cyanobacterial water blooms in Brazil, the increase of eutrophication in our drinking water supplies have permitted abundant growth of cyanobacterial toxic strains in these environments. This work studies the toxin production by a cyanobacterial water bloom which occurred at "Reservatório do Funil", Resende, Rio de Janeiro where Microcystis aeruginosa was the dominant specie. We Verified the occurrence of hepatotoxin in the phytoplanktonic samples and isolated and characterizated the hepatotoxins detected. Toxicity tests were carried out in mouse bioassays. The isolation and characterization of hepatotoxins were performed by isochratic HPLC, FABMS and FABMS/MS analysis. The hepatotoxicity of a water bloom had a positive linear correlation with M. aeruginosa cell percentages. Two hepatotoxin were isolated and characterizated as microcystin-LR and (D-Glu( OCH3)6jmicrocysti~- LR. This is the first time that the latter structure was identified from Microcystis aeruginosa.
Author: Luciana de Paula Lima e Schimidt de Andrade
Title: The influence of high concentrations of zinc in phytoplankton species in Sepetiba Bay, Rio de Janeiro, Brazil.
Year: 1993. Full text (in Portuguese)
Abstract
Sepetiba bay is a water body of fisheries and sonial importance that suffers from metal pollution coming mostly from a zinc and cadmium industry. Several researches have been performed concerning the bay. Three phytoplanktonic species from this water body were studied in laboratory cultures as a contribution to the knowledge of their physiology. Phaeodactylym tricornutum,. Chlorella sp. and Synechocystis sp. were cultivated under increasing zinc concentrations (control= 22µg.L-1; 20Zn=440 µg.L-1; 50Zn= 1100 µg.L-1; 100Zn= 22µg.L-1) and evaluated by means of culture growth (cel1 number • ml- 1), concentration o f chlorophyll a, carbohydrates and proteins. These cultures obtained data were compared taking into account the maximum chlorophyll a and protein concentrations as well as the minimum carbohydrate concentration during the exponential growth phase. The primary production measured with NaH14CO3 (µgC.µgChla-1.h-1) was determined in the exponential phase and the photosynthetic activity was ,calculated. The zinc incorporated to the cells quantified at the end of the cultivation periods by atomic absorption spectrometry. The high zinc levels led to an increase in chlorophyll a synthesis for the three species (up to 85%), and to a and Synechocystis sp. ( 47%). Photosynthetic activity was increased for all contaminated cultures for t he three species, with the exception of 100Zn condition of Chlorella sp,. At the enc of the cultivation periods Zn had been incorporated into the cells of P . triconutum (up to 1400%) and Chlorella sp. (up to 350%) . The conditions 20ZN, 50Zn and 100Zn de Synehocystis sp. incorporated less Zn than the control, indicating a detoxification process for this species. As this three species showed ability to grow even in the mentioned high zinc levels concentrations, and P. tricornutum and Chlorella sp. showed also ability to concentrate this metal, it can be concluded that this is certainly a process that allows zinc to be transferred to other trophic levels in natural environment ..
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