Comunidades de cianobacterias bentónicas, producción y liberación de microcistinas en el río Muga (NE Península Ibérica)

El estudio de las comunidades de cianobacterias en la cuenca del río Muga permite profundizar en la búsqueda de factores que favorezcan o inhiban la acumulación intracelular de microcistinas y su liberación al medio externo en condiciones oligotróficas o con leves signos de eutrofia, al tiempo que permiten aportar nuevos conocimientos que ayuden a mejorar el entendimiento de los mecanismos y procesos que participan en la dinámica de las comunidades de cianobacterias bentónicas y las microcistinas asociadas en ecosistemas fluviales mediterráneos. Se ha realizado un estudio de la estructura y variabilidad de las comunidades de cianobacterias durante un ciclo anual completo mediante muestreos mensuales y muestreos con menor periodicidad durante un segundo ciclo posterior. Dichos muestreos han permitido observar diferentes patrones espacio-temporales a lo largo de todo el período de estudio en los 4 puntos de muestreo seleccionados en la cuenca del río Muga. Estos 4 tramos incluyen 1 punto regulado:aguas abajo del embalse de Boadella, y 3 sin regular:2 cabeceras y un punto aguas abajo de la población de Albanyà. Se han identificado un total de 34 especies de cianobacterias bentónicas en las 4 zonas estudiadas, predominando siempre especies de aguas limpias o poco alteradas. Dos especies, Rivularia biasolettiana (zonas más limpias y expuestas) y Phormidium retzii (sonas esciáfilas o con alta velocidad de corriente), resultan dominantes en gran parte de ellas. Se han reconocido un total de 6 microcistinas acumuladas intracelularmente en la comunidad bentónica de cianobacterias, identificando 3 de ellas: MC-RR, MC-LR y MC-YR. La concentración de microcistinas intracelulares ha disminuído a lo largo del eje principal del río Muga, determinada por diferentes factores físicos, químicos y biológicos. A nivel general, la acumulación total de microcistinas intracelulares ha estado asociada a un aumento de la temperatura del agua, un descenso en la diversidad de la comunidad de cianobacterias y pequeños incrementos de la concentración de fósforo disuelto en el agua. En cuanto a la liberación de microcistinas, ha estado vinculada a distintas perturbaciones soportadas por la comunidad cianobacteriana, tanto asociadas a factores físicos naturales como antrópicos.

Modelling the growth and movement of cyanobacteria in river systems

Bloom-forming and toxin-producing cyanobacteria remain a persistent nuisance across the world. Modelling of cyanobacteria in freshwaters is an important tool for understanding their population dynamics and predicting the location and timing of the bloom events in lakes and rivers. In this article, a new deterministic model is introduced which simulates the growth and movement of cyanobacterial blooms in river systems. The model focuses on the mathematical description of the bloom formation, vertical migration and lateral transport of colonies within river environments by taking into account the four major factors that affect the cyanobacterial bloom formation in freshwaters: light, nutrients, temperature and river flow. The model consists of two sub-models: a vertical migration model with respect to growth of cyanobacteria in relation to light, nutrients and temperature; and a hydraulic model to simulate the horizontal movement of the bloom. This article presents the model algorithms and highlights some important model results. The effects of nutrient limitation, varying illumination and river flow characteristics on cyanobacterial movement are simulated. The results indicate that under high light intensities and in nutrient-rich waters colonies sink further as a result of carbohydrate accumulation in the cells. In turbulent environments, vertical migration is retarded by vertical velocity component generated by turbulent shear stress. (c) 2006 Elsevier B.V. All rights reserved.

Microcystins -LA, -YR, and -LR action on neutrophil migration

Microcystins (MCs) produced by some freshwater cyanobacterial species possess potent liver toxicity as evidenced by acute neutrophil infiltration. Here, we investigate the ability of three structurally distinct toxins (MC-LA, MC-LR, and MC-YR) to evoke neutrophil recruitment per se and their effects on migration pathways. Intravital Microscopic Studies showed that topical application of only MC-LR enhanced the numbers of rolling and adhered leukocytes in the endothelium of postcapillary mesenteric venules. The latter effects may be dependent upon induction of the synthesis and expression Of L-selectin and beta(2)-integrin in neutrophils, as assessed by flow cytometry and RT-PCR, respectively. Conversely, the three toxins promoted direct locomotion of neutrophils and enhanced their migration in response to NO, as measured by Boyden chamber assays, and increased intracellular calcium, a messenger in the chemotaxic process. In conclusion, our results show that MCs act on specific pathways of neutrophil recruitment, indicating their potential effect on neutrophils activation. (C) 2009 Elsevier Inc. All rights reserved.

Functional characterization of the twin ZIP/SLC39 metal transporters, NpunF3111 and NpunF2202 in Nostoc punctiforme

The ZIP family of metal transporters is involved in the transport of Zn2+ and other metal cations from the extracellular environment and/or organelles into the cytoplasm of prokaryotes, eukaryotes and archaeotes. In the present study, we identified twin ZIP transporters, Zip11 (Npun_F3111) and Zip63 (Npun_F2202) encoded within the genome of the filamentous cyanobacterium, Nostoc punctiforme PCC73120. Sequence-based analyses and structural predictions confirmed that these cyanobacterial transporters belong to the SLC39 subfamily of metal transporters. Quantitative real-time (QRT)-PCR analyses suggested that the enzymes encoded by zip11 and zip63 have a broad allocrite range that includes zinc as well as cadmium, cobalt, copper, manganese and nickel. Inactivation of either zip11 or zip63 via insertional mutagenesis in N. punctiforme resulted in reduced expression of both genes, highlighting a possible co-regulation mechanism. Uptake experiments using 65Zn demonstrated that both zip mutants had diminished zinc uptake capacity, with the deletion of zip11 resulting in the greatest overall reduction in 65Zn uptake. Over-expression of Zip11 and Zip63 in an E. coli mutant strain (ZupT736::kan) restored divalent metal cation uptake, providing further evidence that these transporters are involved in Zn uptake in N. punctiforme. Our findings show the functional role of these twin metal uptake transporters in N. punctiforme, which are independently expressed in the presence of an array of metals. Both Zip11 and Zip63 are required for the maintenance of homeostatic levels of intracellular zinc N. punctiforme, although Zip11 appears to be the primary zinc transporter in this cyanobacterium, both ZIP's may be part of a larger metal uptake system with shared regulatory elements.

Characterization of two cation diffusion facilitators NpunF0707 and NpunF1794 in Nostoc punctiforme

AIMS: To characterize genes involved in maintaining homeostatic levels of zinc in the cyanobacterium Nostoc punctiforme. METHODS AND RESULTS: Metal efflux transporters play a central role in maintaining homeostatic levels of trace elements such as zinc. Sequence analyses of the N. punctiforme genome identified two potential cation diffusion facilitator (CDF) metal efflux transporters, Npun_F0707 (Cdf31) and Npun_F1794 (Cdf33). Deletion of either Cdf31or Cdf33 resulted in increased zinc retention over 3 h. Interestingly, Cdf31(-) and Cdf33(-) mutants showed no change in sensitivity to zinc exposure in comparison with the wild type, suggesting some compensatory capacity for the loss of each other. Using qRT-PCR, a possible interaction was observed between the two cdf's, where the Cdf31(-) mutant had a more profound effect on cdf33 expression than Cdf33(-) did on cdf31. Over-expression of Cdf31 and Cdf33 in ZntA(-) - and ZitB(-) -deficient Escherichia coli revealed function similarities between the ZntA and ZitB of E. coli and the cyanobacterial transporters. CONCLUSIONS: The data presented shed light on the function of two important transporters that regulate zinc homeostasis in N. punctiforme. SIGNIFICANCE AND IMPACT OF THE STUDY: This study shows for the first time the functional characterization of two cyanobacterial zinc efflux proteins belonging to the CDF family.

Estrutura e dinâmica do fitoplânctone bacterioplâncton em cultivos de camarão no Rio Grande do Norte-Brasil: impacto sobre o ambiente natural

This work aimed to study the structure and dynamic of Phytoplankton and Bacterioplankton in a complete cycle of shrimp cultivation (Litopenaeus vannamei) and determine the environmental factors responsible for the structural changes of these communities. The study was realized in a saltwater shrimp farm (Macaíba, RN), between September/2005 and February/2006, and in a freshwater shrimp farm (Ceará Mirim, RN), between May/2007 and September 2007. The samplings were collected weekly in saltwater farm and every fifteen days in freshwater farm. Total phosphorus, chlorophyll a and environmental parameters (pH, dissolved oxygen, salinity, temperature, depth and water transparency) were measured. Qualitative and quantitative analysis of the phytoplankton and bacterioplankton were carried out. The Shannon-Wiener ecologic indexes of diversity and the Pielou equitability indexes were calculated to the phytoplankton. Bacterial density was determined by epifluorescence microscopy. The data were statistically analyzed by Pearson correlation and t-Test. Chlorophycea were predominat in salt water and in the captation/drainage point (24 to 99%). Diatoms had higher wealth. The species Choricystis minor had the highest occurrence (100%) and dominance (90-100%), thus showing its adaptation to the high temperatures, salinity and low water transparency conditions. Filamentous Cyanobacteria like Oscillatoria sp., Pseudoanabaena sp. and Phormidium sp. had constant levels. The negative correlation between chlorophycea and water transparency, and the positive correlation between chlorophyll a and salinity, showed that the phytoplankton was well adapted to the low transparency and to the high salinity. The bacterioplankton was negatively correlated with the total phosphorus and salinity. In freshwater, Cyanobacteria were predominant (>80%), presenting some producers of toxins species like Microcystis sp., Aphanizomenon sp., Cylindrospermopsis raciborskii e Anabaena circinalis. Cyanobacterial density and total phosphorus and chlorophyll a concentrations exceeded the maximum value allowed by legislation. The means of total phosphorus varied from 264 to 627 Wg.L-1 and the means of chlorophyll a oscillated between 22 and 182 Wg.L-1. The phytoplankton species were selected by low availability of the light, high pH, temperature and high availability of total phosphorus. The bacterioplankton showed high densities (5,13 x 107 to 8,50 x107 Bac.mL-1). The studied environments (ponds and rivers) presented a high level of trophic state based on the high concentrations of chlorophyll a and total phosphorus and cyanobacteria dominance. The composition of species in the ponds and rivers was similar, as well as high concentrations of total phosphorus and chlorophyll a, highlighting the pollution caused by the discharges of the farms in natural environment

Avaliação da toxicidade de florações naturais e de cultura de cianobactérias: efeitos sobre Ceriodaphnia silvestri (Crustacea, Cladocera)

The incidence of toxic cyanobacterial blooms is one of the important consequences of eutrophication in aquatic ecosystems. It is a very common phenomenon in reservoirs and shrimp ponds in the State of Rio Grande do Norte (RN), Brazil. Cyanobacterias produce toxins which can affect aquatic organisms and men trough the food chain. Aiming to contribute to the studies of cyanobacterias in RN, we propose: a) to evaluate the toxicity of isolated cyanobacterias in important fresh-water environments; and b) to verify the effects of both natural and cultured blooms occurred in reservoirs for human supply and in the cladoceran Ceriodaphnia silvestrii. This study was carried out using samples of natural blooms occurred between March and October of 2004 in Gargalheiras Dam (08º L e 39º W), in July of 2004 in Armando Ribeiro Gonçalves Dam (06o S e 37o W) and in commercial shrimp ponds (Litopenaeus vannamei) located in fresh-water environments. The samples were collected with plankton net (20µm.) for identification, isolation and obtaining of phytoplanktonic biomass for liophilization and later toxicity bioassays. The toxicity of cultured samples and natural blooms was investigated through bioassays in Swiss mice. Quantification of cyanobacteria in samples was conducted following the Ütermol method, with 300mL samples fixed with lugol. The toxicity test with Ceriodaphnia silvestrii followed ABNT, 2001 recommendations, and were accomplished with natural hepatotoxic bloom s samples and cultured samples of both non-toxic and neurotoxic C. raciborskii. In this test, five newborns, aged between 6 and 24 hours, were exposed to different concentrations (0 a 800 mg.L-1) of crude cyanobacterial extracts during 24 and 48 hours. Three replicates were used per treatment. The pH, temperature and dissolved oxygen at the beginning and after 24 and 48hours from the test were measured. We estimated the CL50 through the Trimmed Spearman-Karber method. The blooms were constituted by Microcystis panniformis, M. aeruginosa, Anabaena circinalis, Cylindrospermopsis raciborskii and Planktothrix agardhii, producers of mycrocistin-LR confirmed with HPLC analysis. Samples of hepatotoxic blooms registered toxinogenic potential for C. silvestrii, with CL50-24h value of 47.48 mg.L-1 and CL5048h of 38.15 mg.L-1 for GARG samples in march/2005; CL50-24h of 113,13 mg.L-1 and CL5048h of 88,24 mg.L-1 for ARG July/2004; CL50-24h of 300.39 mg.L-1 and CL50-48h of 149.89 mg.L-1 for GARG October/2005. For cultured samples, values of CL50-24h and CL50-48h for C. raciborskii toxic strains were 228.05 and 120.28 mg.L-1, respectively. There was no mortality of C. silvestrii during the tests with non-toxic C. raciborskii strain. The toxicity test with C. silvestrii presented good sensitivity degree to cyanotoxins. The toxicity of natural hepatotoxic blooms samples (microcystins) and cultured neurotoxic saxitoxins producer samples analyzed in this study give us strong indications of that toxin s influence on the zooplanktonic community structure in tropical aquatic environments. Eleven cyanobacteria strains were isolated, representing 6 species: Anabaenopsis sp., Cylindrospermopsis raciborskii, Chroococcus sp., Microcystis panniformis, Geitlerinema unigranulatum e Planktothrix agardhii. None presented toxicity in Swiss mice. The strains were catalogued and deposited in the Laboratório de Ecologia e Toxicologia de Organismos Aquáticos (LETMA), in UFRN, and will be utilized in ecotoxicológical and ecophysiological studies, aiming to clarify the causes and control of cyanobacterial blooms in aquatic environments in RN. This state s reservoirs must receive broader attention from the authorities, considering the constant blooms occurring in waters used for human consumption

Avaliação dos efeitos de um extrato bruto de cianobactérias contendo microcistinas sobre o desempenho reprodutivo e a embriofetotoxicidade de ratos

The presence of cyanobacterial blooms in reservoirs intended for supply to the population can create public health problems for many species could produce potentially toxic compounds and these are not eliminated in the conventional procedures used in water treatment plants. So even in amounts less than the maximum allowable limit imposed by MS, cyanotoxins can be present in drinking water distributed to the population, creating a chronic exposure. There is little information about the long-term effects of oral exposure to cyanotoxins. This work aimed to show the exposure orally (v.o) of animals to a crude extract of cyanobacteria containing cyanotoxins to evaluate the reproductive performance of pregnant rats and their offspring and fertility of male rats. The presence of microcystins (MCs) in samples collected during the flowering processes in freshwater reservoirs in the Rio Grande do Norte, was analyzed by enzyme immunoassay and its variants have been identified and quantified by chromatographic methods. It was observed that by administration v.o. cyanobacterial extract containing MCs (40, 100 or 250 ng of MCs / kg / day) did not cause systemic toxicity in adult rats or effect on reproductive performance of male and female rats treated. It was also not observed any changes in skeletal study in the offspring of pregnant rats treated with the extract above. Because the solutions used contained MCs in a concentration equal to or greater than the tolerable daily intake for MCs, the results suggest, therefore, that the development of this work contributed to better assess public health risk as the oral exposure to cyanotoxins, increasing thus the credibility of the maximum allowable limit (LMP) of MCs in drinking water distributed to the population of several countries that use the LMP established by WHO in its legislation

Efeitos de florações de cianobactérias sobre os cladóceros ceriodaphnia dubia e daphnia gessneri

Cyanobacterial blooms are common in eutrophic reservoirs in Brazilian northeastern semi-arid. Given this reality, the present study aimed to analyze the effect of potentially toxic cyanobacterial blooms in Gargalheiras reservoir (semi-arid) on the cladocerans Ceriodaphnia dubia and Daphnia gessneri. In vitro chronic bioassays were performed with reservoir water dilutions from August/2011 to May/2012 and the following effects were evaluated on: intrinsic rate of population growth (r), reproductive parameters (age of first reproduction and fecundity per capita) and cladocerans movements. Phytoplankton was dominated by Cylindrospermopsis raciborskii and Planktothrix agardhii and saxitoxin and microcystin were detected in reservoir water. In most months C. dubia showed differences in r between control (absence of cyanobacteria) and treatments, and has shown negative effects on reproductive parameters. In all months paralysis of swimming movements was observed in C. dubia when both C. raciborskii and saxitoxin (cyanotoxin neurotoxic) were present in water. While C. dubia was sensitive to the reservoir water containing cyanobacteria, D. gessneri showed less intense negative effects in r and reproductive parameters. Furthermore, D. gessneri showed no paralysis of swimming movements. These results support the hypothesis in the literature that D. gessneri is resistant to the Cylindrospermopsis effects. The clone‟s life history may be a key to understand the results. The C. dubia clone, isolated from eutrophicated environment, is in the lab for ten years and it is an exotic species in Brazil. D. gessneri is a common species in the country and this clone was isolated from the Gargalheiras reservoir (where there are constant blooms of potentially toxic cyanobacteria) a year ago. Perhaps the recent contact with cyanobacteria explain the higher resistance presented by this D. gessneri clone. In conclusion, the cladocerans studied have different levels of sensitivity to cyanobacteria, characterizing species-specific responses

O efeito do regime hidrológico do semiárido na composição de espécies durante dominância de cianobactérias em um reservatório tropical

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

A lectin from the lichenized Basidiomycete Dictyonema glabratum

A lectin with hemagglutinating activity has been isolated from an aqueous extract of the symbiotic phenotype of Dictyonema glabratum and its cyanobacterial photobiont Scytonema sp. The purified lectin had a pi of 6.8 and its molecular mass was investigated by electrospray ionization mass spectrometry, gel filtration and SDS-PAGE, which indicated its native conformation as a dimer formed by two identical subunits of 16540 Da. The lectin is a glycoprotein with a low degree of glycosylation, containing galactose, xylose, glucose and mannose as neutral monosaccharides, in addition to glucosamine, which could indicate both N- and O-linkages. Amino acid analysis showed the predominance of nonpolar residues such as phenylalanine. Agglutination of human erythrocytes required divalent cations, which is affected by addition of EDTA. The lectin was more stable at 30 degreesC or less for at least 1 h and between pH 5.0 and 7.0. Among the various compounds tested for hemagglutination inhibition, N-acetylgalactosamine was the most active. The potential role of this lectin in recognition of the compatible cyanobacterial photobiont is discussed.

Avaliação do potencial de toxicidade de florações de cianobactérias como subsídio para ações de gestão ambiental em açudes do semiárido potiguar

Blooms negatively compromise the aquatic environment, plants, animals and human health.Some species are toxin-producing, such as the neurotoxin saxitoxin and the hepatotoxin microcystin, which may contaminate water reservoirs, as those existing in the semiarid region of Rio Grande do Norte (Brazil) which are used to supply the population, fishing, aquiculture and recreational activities, thereby providing the risk of human exposure through water intake, dermal contact and respiratory tract. Thus, it is recommended a constant monitoring of the density of cyanobacteria with the quantification of cyanotoxins. One goal with this work was the monitoring of water in four reservoirs in semiarid of RN through the identification and enumeration of cyanobacteria and through the identification and quantification of cyanotoxins by ELISA. Furthermore, we intended to assess the environmental perception of farmers and artisanal fishers in reservoirs of semiarid of RN through semi-structured interviews with questions mostly related to water and eutrophication. Through these objectives the aim was the development of management strategies for aquaculture and prevention of risks to public health. The results showed that the highest values of microcystins were found in the rainy season. Standards for drinking water, according to the guidelines of Ministry of Health 2914/2011 and CONAMA 357/05, setting the maximum values for raw water density of cyanobacteria: 50,000 cel.mL-1; microcystin: 1 μg. L-1 and saxitoxin: 3 μg. L-1. The values found for microcystin ranged between 0.00227 μg. L-1 and 24.1954 μg. L-1. From 128 samples analyzed, 27% were above the limit. There was no clear seasonal pattern for saxitoxins and their values ranged between 0.003 μg. L-1 and 0.766 μg. L-1 with none of the values above the limit. Furthermore, 76% of the densities of cyanobacteria values were above the limit. About environmental perception, 52 interviews were conducted and the results show that the respondents recognize the main uses of water of the rervoirs, recognize the importance and have a positive view about the reservoirs. They also realize that the water has a poor quality and can cause health problems. The results provide data showing the persistence of cyanobacteria and cyanotoxins, many times over the limit, reinforcing the importance of constant monitoring. The assessment of environmental perception gives foundation for later proposed environmental education linked to public health management into the context of this particular population, making it more effective

New aerophytic morphospecies of Nostoc (Cyanobacteria) from São Paulo State, Brazil

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

New aerophytic morphospecies of Cyanobacteria from tropical forest fragments in northwestern São Paulo state, Brazil

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Algas e cianobactérias aerofíticas de fragmentos de floresta estacional semidecidual no noroeste paulista

Pós-graduação em Ciências Biológicas (Biologia Vegetal) - IBRC