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Caracterización morfológica y molecular de cianobacterias filamentosas aisladas de florecimientos de tres lagos urbanos eutróficos de la ciudad de México
Abstract
Cyanobacteria are traditionally classified based on morphological and ecological traits, which can result in incorrect identifications due to phenotypic plasticity, and in the case of isolated strains, due to morphological changes provoked by the culture conditions. The gene 16S rRNA is useful for taxonomic identification, helping to overcome the aforementioned problems. Using morphological and molecular characteristics, we isolated and identified filamentous cyanobacteria present in blooms in three eutrophic urban lakes in Mexico City with the aim of recognizing genera of cyanobacteria other than Microcystis that may be present. Water samples were collected during March 2007, and through micromanipulation we obtained monospecific cultures that were preliminarily identified using morphological traits. In addition, total DNA was extracted, and a partial region of the 16S rRNA gene was amplified to corroborate the taxonomic identifications; these were confirmed through their phylogenetic positioning. In this way most of the isolated strains identified by their morphological traits could be confirmed at the generic level. We identified strains of Anabaenopsis, Arthrospira, Leptolyngbya, Limnothrix, Geitlerinema, Phormidium, Planktolyngbya, Planktothrix, Pseudanabaena and Spirulina, showing the diversity of cyanobacteria that could be recorded in blooms even though the dominant taxa could be Microcystis spp. To accurately determine taxonomic identity we strongly recommend the use of both techniques in order to avoid errors due to phenotypic variation provoked by environmental factors or culture conditions.
... En la zona urbana de Xochimilco, Ciudad de México, específicamente en las aguas de la Pista Olímpica de Remo y Canotaje Virgilio Uribe, existe un problema ambiental desde hace varias décadas, debido a la presencia y permanencia de florecimientos derivados del crecimiento excesivo de Microcystis (Pineda-Mendoza et al. 2011). ...
... Microcystis fue recolectada en abril de 2017 en la Pista Olímpica (19º 16' 20.58" N,99º 6' 16.72" O) de la Ciudad de México. Este cuerpo de agua mide aproximadamente 2200 m de largo, 125 m de ancho, tiene 2 m de profundidad y mantiene su volumen mediante aporte pluvial y vertido de aguas tratadas (Pineda-Mendoza et al. 2011). ...
... del crecimiento excesivo de Microcystis(Pineda-Mendoza et al. 2011). En esta región, los FAN de cianobacterias no han recibido la suficiente atención y, en general, se ignoran los riesgos que representan para la salud humana y la biota. ...
El crecimiento de diversas cianobacterias nocivas (Microcystis) se ha intensificado a nivel mundial como resultado de numerosas y rápidas alteraciones ambientales. En este estudio la cianobacteria Microcystis fue aislada de la Pista Olímpica de Remo y Canotaje Virgilio Uribe de la Ciudad de México y se investigaron: 1) la influencia del aumento de nitrógeno (N) y fósforo (P) en el crecimiento de Microcystis y 2) la toxigenicidad potencial de Microcystis mediante bioensayos de mortalidad de Artemia sp. En 2017, dos cepas clonales de Microcystis (Mc17C2 y Mc17C3) se establecieron en condiciones controladas de laboratorio. Dichas cepas crecieron en una cámara iluminada con irradianza de 19.6 µmol/m2/s con un fotoperiodo de 12 h luz:12 h oscuridad a temperatura ambiente (~ 24 ± 2 ºC). Se cultivó Artemia sp. y se seleccionaron juveniles del crustáceo para alimentarlos con diferentes concentraciones de Microcystis. El aumento en la concentración de N y P se asoció con mayores tasas de crecimiento de Microcystis; sin embargo, su crecimiento óptimo no coincidió con los niveles más altos de nutriente en los cultivos. La ingestión de Microcystis por juveniles de Artemia causó mortalidad del 50 al 100 % en un lapso de 48-72 h cuando la abundancia fluctuó entre 3 × 105 y 3 × 106 cel/mL. En general, Artemia sp. no mostró sensibilidad al consumo de Microcystis en las primeras 24 h. Se concluye que el incremento y la disponibilidad de nutrientes disueltos conducen a mayores tasas de crecimiento de Microcystis con efecto potencial sobre el zooplancton.
... Root DNA was extracted with the enzymatic lysis technique of Sambrook and Russell 63 . The extraction of DNA from the seeds was based mainly on a protocol proposed by CIMMYT 66 , but with some adaptations based on the method described by Pineda et al. 67 . First, tissues were disinfected and then macerated with liquid nitrogen to obtain a fine powder. ...
... Nine mL of extraction solution (Tris 1 M, EDTA 0.5 M, CTAB, NaCl 5 M, BME 14 M) was added and incubated at 70 °C for 1 h 66 . Proteins were precipitated with a solution of chloroform/isoamyl alcohol (24:1) 67 . Phenol was used for a higher purity extraction followed by precipitation with ethanol and washed to purify DNA 66 . ...
Rhizosphere and root endophytic bacteria are crucial for plant development, but the question remains if their composition is similar and how environmental conditions, such as water content, affect their resemblance. Ricinus communis L., a highly drought resistant plant, was used to study how varying soil water content affected the bacterial community in uncultivated, non-rhizosphere and rhizosphere soil, and in its roots. Additionally, the bacterial community structure was determined in the seeds of R. communis at the onset of the experiment. Plants were cultivated in soil at three different watering regimes, i.e. 50% water holding capacity (WHC) or adjusted to 50% WHC every two weeks or every month. Reducing the soil water content strongly reduced plant and root dry biomass and plant development, but had little effect on the bacterial community structure. The bacterial community structure was affected significantly by cultivation of R. communis and showed large variations over time. After 6 months, the root endophytic bacterial community resembled that in the seeds more than in the rhizosphere. It was found that water content had only a limited effect on the bacterial community structure and the different bacterial groups, but R. communis affected the bacterial community profoundly.
... Se ha reportado a especies de este género en el Sur de Francia (Chomérat et al., 2007), México (Komárek & Komárková, 2002;Novelo, 2011;Pineda et al., 2011), Perú (Saavedra, 2015, en Uruguay (Pérez, 2002;Ferrari et al., 2011) y en Argentina (Wenzel & Díaz, 2008). La forma de las células es rectangular, cuyo tamaño puede variar de 3 a 6 µm de largo y 1 a 2 µm de ancho. ...
... Se ha registrado la presencia de Phormidium en Sur de Francia (Chomérat et al., 2007), México (Novelo, 2011;Pineda et al., 2011), Honduras (Hernández & Marin, 2017), Brasil (Nishimura et al., 2014, ) y Argentina (Huber et al., 2011;Galea et al., 2014). En Paraguay, esta especie ha sido registrada en la Cuenca del Arroyo Tacuary, Departamento de Itapúa (López Pereira, 2013) y en el Lago Ypacaraí (Delgado et al., 2014). ...
En Paraguay, son pocos los científicos que se dedican a la investigación de la composición Ficológica de los sistemas de agua del territorio nacional. En consecuencia, se genera un impacto negativo sobre los resultados de análisis en monitoreos de gestión ambiental y posibilidades de aplicación en áreas de Paleolimnología, Ecología y Biotecnología. En cierto aspecto, la falta de información presenta una ventaja para quien esté atraído por estudiar la riqueza ficológica del Paraguay debido a que se encuentra ante grandes áreas potenciales de investigación y desarrollo de servicios y productos. Sin embargo la falta de información siempre es una desventaja en especial a la hora de analizar las problemáticas ambientales e intentar soluciones para ellas. Por ello, es importante comenzar a revertir el panorama de desconocimiento sobre las algas en Paraguay. Si bien, todavía son pocos los trabajos publicados sobre especies ficológicas, son muy importantes como base o punto de partida.
Este humilde catálogo, pretende ser una herramienta científica y
académica así como una base importante para el conocimiento de un
grupo ficológico importante . Cuando hablamos de recursos ficológicos, nos referimos a cualquiera o a todos los grupos de algas tanto macroscópicos como microscópicos. Este material se constituye como el primer material que hace una colaboración específica en el conocimiento del grupo de Cyanophytas, algas microscópicas verdeazuladas. Habitualmente pueden ser encontradas formando parte de la flora microscópica superficial de sistemas de agua naturales o artificiales. El área de estudio para
este trabajo fue el estanque artificial del campus de la UNA, un pequeño espacio aledaño a las instalaciones de la Facultad de Ciencias Exactas y Naturales, donde las especies de Cyanophytas fueron identificadas y hoy enriquecen la base de datos de especies de algas identificadas para Paraguay.
El catálogo provee una lista de 28 especies de cianófitas identificadas. También se encuentra la descripción de cada especie con la fotomicrografía correspondiente. Como complemento muy innovador, en este tipo de trabajos bibliográficos, para algunas de las especies se incluyeron un pequeño resumen de artículos científicos que relatan avances biotecnológicos logrados con algunas de las especies de este grupo, así como en distintos sectores de producción o de servicios.
El trabajo se complementa ofreciendo información con algunos
procedimientos y metodologías para trabajar con este u otro grupo de microalgas, en campo y laboratorio. Una clave de identificación bien detallada por género de las cianófitas.
... A total of 17 strains of filamentous cyanobacteria isolated in March 2007 from three urban lakes in Mexico City were selected for this study. The lakes are located at Chapultepec Park First Section (19º 25' 25.90" N, 99º 11' 5.03" W), Alameda Oriente Ecological Park (divided into five ponds, 19º 26' 13.09" -19º 26' 8.36" N and 99º 3' 15.36" -99º 3' 22.19" W) and the Olympic Rowing-Canoeing Course "Virgilio Uribe" at Cuemanco (19º 16' 20.58" N, 99º 6' 16.72" W) (Pineda-Mendoza et al., 2011). These strains were taxonomically identified based on their morphological characteristics and the genetic marker 16S rRNA. ...
... A total of 17 cyanobacteria strains that had been previously identified by partial 16S rRNA gene sequencing were examined (Table 1), clustering into 10 different genera (Pineda-Mendoza et al., 2011). ...
Cyanobacterial blooms are of great importance because of the toxic effects that these microorganisms are able to induce, particularly on aquatic organisms. Microcystins (MCs) are the principal toxins biosynthesized by cyanobacteria and are powerful inhibitors of the protein phosphatases 1 and 2A. Zooplankton filter feeders such as cladocerans are directly affected by MCs as a result of ingestion of cyanobacteria or contact with intracellular products when cyanobacterial cells break up during and after blooms. A total of 17 strains of filamentous cyanobacteria isolated from three urban lakes in Mexico City were characterized using the microcystin synthetase region mcyA-Cd. Acute 48-h toxicity was evaluated in different strains using the cladoceran Daphnia magna and total microcystin content was determined by enzyme-linked immunosorbent assay (ELISA). The mcyA-Cd region was amplified in 16 microcystin-producing strains; microcystins were detected in eight strains with values ranging from 0.1422 to 2.772 μg L-1. Nevertheless, all aqueous crude extracts induced acute toxicity in D. magna with LC50 values from 363.91 to 741.8 mg L-1 (dry weight). The toxicity observed in non-microcystin-producing strains may be induced by cyclic peptides other than microcystins (anabaenopeptins, microviridins and cyclamides). The results obtained warn of the toxigenic potential of filamentous cyanobacteria, since though Microcystis spp. is frequently predominant in blooms, other toxins and intracellular metabolites released by filamentous cyanobacteria may induce toxicity on aquatic organisms as well as humans.
... A total of 17 strains of filamentous cyanobacteria isolated in March 2007 from three urban lakes in Mexico City were selected for this study. The lakes are located at Chapultepec Park First Section (19º 25' 25.90" N, 99º 11' 5.03" W), Alameda Oriente Ecological Park (divided into five ponds, 19º 26' 13.09" -19º 26' 8.36" N and 99º 3' 15.36" -99º 3' 22.19" W) and the Olympic Rowing-Canoeing Course "Virgilio Uribe" at Cuemanco (19º 16' 20.58" N, 99º 6' 16.72" W) (Pineda-Mendoza et al., 2011). These strains were taxonomically identified based on their morphological characteristics and the genetic marker 16S rRNA. ...
... A total of 17 cyanobacteria strains that had been previously identified by partial 16S rRNA gene sequencing were examined (Table 1), clustering into 10 different genera (Pineda-Mendoza et al., 2011). ...
Cyanobacterial blooms are of great importance because of the toxic effects that these microorganisms are able to induce, particularly on aquatic organisms. Microcystins (MCs) are the principal toxins biosynthesized by cyanobacteria and are powerful inhibitors of the protein phosphatases 1 and 2A. Zooplankton filter feeders such as cladocerans are directly affected by MCs as a result of ingestion of cyanobacteria or contact with intracellular products when cyanobacterial cells break up during and after blooms. A total of 17 strains of filamentous cyanobacteria isolated from three urban lakes in Mexico City were characterized using the microcystin synthetase region mcyA-Cd. Acute 48-h toxicity was evaluated in different strains using the cladoceran Daphnia magna and total microcystin content was determined by enzyme-linked immunosorbent assay (ELISA). The mcyA-Cd region was amplified in 16 microcystin-producing strains; microcystins were detected in eight strains with values ranging from 0.1422 to 2.772 µg L-1. Nevertheless, all aqueous crude extracts induced acute toxicity in D. magna with LC50 values from 363.91 to 741.8 mg L-1 (dry weight). The toxicity observed in non-microcystin-producing strains may be induced by cyclic peptides other than microcystins (anabaenopeptins, microviridins and cyclamides). The results obtained warn of the toxigenic potential of filamentous cyanobacteria, since though Microcystis spp. is frequently predominant in blooms, other toxins and intracellular metabolites released by filamentous cyanobacteria may induce toxicity on aquatic organisms as well as humans.
... In Mexico, P. agardhii has been previously reported in hypereutrophic lakes. Komárek and Komárková-Legnerová [17] and Pineda-Mendoza et al. [18] analyzed the taxonomic and molecular characteristics of P. agardhii in Xochimilco and Chapultepec urban lakes in Mexico City. Likewise, Vasconcelos et al. [19] reported this species in the Valle de Bravo dam and detected MCs using ELISA. ...
The development of anthropic activities during recent years has led to an increase in nutrient fluxes in the Río Grande de Comitán and Montebello Lakes National Park, Mexico. In turn, this has modified the dynamics of the biotic community, specifically favoring the presence of cyanobacteria tolerant to contamination. The continual and massive presence of Planktothrix species (spp.) in the system suggests a potential detrimental impact for economic issues and human health. In this study, we identify the morphological and molecular characteristics of Planktothrix populations from seven tropical (1,380–1,740 masl, 23.0–25.5°C) and calcareous lakes and two ponds from a water treatment plant. We also assess the ecological drivers that could be related to the presence of cyanotoxins in the system. The ecological preferences, morphology, 16S rRNA structure, and 16S-23S rRNA internal transcribed spacer found evidence for three species: P. agardhii distributed in neutral to slightly basic water (pH = 7.7–8.7), and P. spiroides and Planktothrix sp. in alkaline waters (pH = 9.1). The presence of the mcyE gene and its validation by liquid chromatography confirmed the presence of two microcystin variants (MC-RR and MC-LR) in at least three populations of P. agardhii. These microcystins put the health of the ecosystem and its inhabitants at risk, a condition that should be addressed and resolved with a water management and detoxification strategy in the basin.
... Tradicionalmente, las cianobacterias se han clasificado con base en caracteres morfológicos como: el ancho, la forma y disposición de los tricomas; el tamaño de las células; los planos de división; la pigmentación; y la presencia de caracteres como vacuolas de gas y vaina (Baker, 1992;Valerio et al., 2009). Sin embargo, debido a la flexibilidad de adaptación de las cianobacterias, estas pueden llegar a presentar cambios fisiológicos debido a las condiciones del clima, hábitat, disponibilidad de recursos, competencia por los mismos e inclusive en condiciones controladas de laboratorio (Pineda-Mendoza et al., 2011). Por tal motivo, se ha evidenciado la necesidad de complementar la identificación tradicional con técnicas de biología molecular, debido a que el genoma no se ve afectado en la misma proporción por factores externos (Al-Haj et al., 2016). ...
El presente trabajo colectó, aisló, identificó y caracterizó el potencial antimicrobiano de dos especies filamentosas de cianobacterias de la Amazonia colombiana. Se utilizaron técnicas microbianas de rayado en medio sólido y dilución para obtener las cepas con una sola especie. La identificación se realizó por caracteres morfológicos y secuenciado de la sección conservada Cya de gen 16s. Se identificaron por reacción en cadena de la polimerasa (PCR) la presencia de genes que secuencian policétidos sintasas (PKS) y péptidos no ribosomales sintetasas (NRPS) asociados a rutas productoras de metabolitos secundarios. Se evaluó la actividad antimicrobiana de extractos polares y no polares con una batería Gram positiva, dos Gram negativas y una cepa fúngica. Se identificó la cepa 5 como Limnothrix vacuolifera (Skuja) Komárek y la cepa 9 como Lymnothix planktonica (Wołoszyńska) Meffert. En las dos cepas se encontraron PKS y NRPS. Finalmente, los extractos polares inhibieron el crecimiento de Enterococcus faecalis, Escherichia coli y Klebsiella pneumoniae y una cepa Penicillium sp. Los extractos no polares no mostraron actividad antimicrobiana. Es el primer registro de estas para la Amazonia colombiana y el primer registro de actividad antibiótica de cianobacterias continentales colombianas.
... Additionally, blue spaces were, and still are, used for residential and industrial wastewater disposal [21,22]. Consequently, many of them are currently neglected, eutrophicated, and polluted [23][24][25]. ...
Lacustrine blue spaces provide benefits to the urbanites and wildlife habitat. Their availability varies depending on the city in which they are established and intra-urban social interactions. We analyzed the presence, distribution, and size of lentic water bodies in Mexico’s 145 most populated cities. We searched for patterns in their distribution concerning demographic, socioeconomic, and geographic data, aiming to understand their socio-ecological interactions in cities. We digitized lacustrine spaces to obtain their number per city, total surface, area of blue space per inhabitant, and surface as a percentage of the city’s total area. We tested for relationships between their number and surface and city population, hydrological regions, and urban marginalization index through linear and generalized linear models. We delimited 1834 lacustrine blue spaces, finding almost two-thirds of them artificial. Their presence and surface in Mexican cities were generally low, except for hydrological regions close to the Gulf of Mexico. Their number and surface decreased as the urban marginalization index increased. The lack of equitable provision of lacustrine space at the national level has implications for urban planning and land management. Blue spaces should maximize their ecosystem services’ provision for the whole society to promote cities’ sustainability and resilience.
... Después de ser observadas en el microscopio, los microorganismos que presentaron características morfológicas de cianobacterias como el color, la forma, el tamaño de las células, la presencia o ausencia de filamentos, número de tricomas y la presencia, forma y tamaño de heterocistos (Pineda-Mendoza et al., 2011), fueron seleccionados para ser asilados mediante la técnica de medios de cultivo líquidos selectivos (Tenorio-García-Blásquez et al., 2018), para la cual se utilizaron los medios BG11 y BG11 0 (Rippka et al., 1979), que son específicos para cianobacterias con la diferencia de que el segundo no contiene nitrógeno y solo las cianobacterias capaces de fijarlo se pueden desarrollar en este medio. Su composición se muestra en la tabla 1. ...
Resumen Las cianobacterias comprenden un gran y diverso grupo de microorganismos procariotas cuyo origen se remonta hace 3.5 millones de años. Son las principales productoras del oxígeno de nuestro planeta, forman la base de las redes tróficas, tienen una amplia distribución en cualquier tipo de ecosistema terrestre o acuático, entre ellos los ambientes con altas temperaturas y son fuente de una amplia gama de productos de alto valor agregado, los cuales han despertado el interés dentro del campo de la biotecnología actual, de tal manera que se han desarrollado aplicaciones en muchas áreas, siendo la agricultura una de las más importantes. Con base a lo anterior, se estableció como objetivo de este avance de investigación, la exploración de las aguas termales del brote de San Diego de Alcalá, Aldama, Chihuahua, en búsqueda de cianobacterias con potencial uso biotecnológico en el campo chihuahuense. Los resultados evidenciaron la presencia de cianobacterias que pueden ser empleadas en el desarrollo de aplicaciones biotecnológicas, lo cual representa una alternativa viable para el desarrollo del campo chihuahuense.
... The level of urbanization is another essential factor impacting house prices, with city house prices usually higher than those in rural areas. The income prospects for residents is another factor that affects their desire to buy a house [47,48]. Government policies, such as rent-to-own models and stricter eligibility criteria for homebuyers, can help regulate housing prices to some extent [49,50]. ...
With the rapid development of digital finance, the implementation of digital infrastructure is becoming increasingly significant. Broadband construction is an important part of the communication network, and can promote urban infrastructure in cities. However, whether the development of broadband can affect housing prices by affecting the urban infrastructure and the convenience of residents is a question worth addressing. In this study, using panel data regarding cities in China, we used the spatial multi-period difference-in-differences (SDID) model to investigate the utility of the Broadband China Policy (BCP) on urban house prices and the mechanism of impact. We found that the BCP can increase house prices, and that this impact has a positive spatial spillover effect. This conclusion still held after a series of tests such as parallel trend tests, placebo tests, and the exclusion of other policy effects. The BCP can increase house prices by improving urban infrastructure, promoting urbanization, and optimizing urban industrial structure. In addition, we conducted a heterogeneity analysis by taking into consideration the administrative level, economic development level, and location of cities. The findings of this paper not only enrich the research on the BCP and housing prices, they also provide policy recommendations in terms of urban land use and sustainable development.
... Collected roots were washed with sterile water, disinfected with 10% hydrogen peroxide, 2.5% sodium hypochlorite, soaked in ethanol and rinsed with sterile water. Metagenomic DNA was extracted using a mechanical (CIMMYT, 2006;Pineda-Mendoza et al., 2011) and enzymatic lysis method (Sambrook et al., 1989). The DNA extracted with the different methods was pooled and used for downstream analyses. ...
Rhizosphere and endophytic microbial communities are crucial for plant fitness and affect how plants cope with abiotic stress. In this study, we provide evidence that drought stress affected alpha and beta diversity of fungal communities associated with the roots and rhizosphere of castor bean (Ricinus communis) through metabarcoding of 18S rRNA gene. Plants were cultivated in soil columns in the greenhouse at three different watering regimes, i.e., 50% water holding capacity (WHC; wet) or adjusted to 50% WHC every 2 weeks (dry) or every month (extremely dry). Ascomycota, Basidiomycota, Chytridiomycota and Fusarium dominated the soil and rhizosphere and Fusarium, Cladosporium, Mucor, Cystofilobasidium, Penicillium and Malassezia the roots. Under extremely dry conditions, root and rhizosphere taxonomic and functional alpha diversity increased compared to the wet treatment. However, the species turnover decreased in the stressed compared to the non-stressed roots, enriching specific fungal groups. Drought did not affect the association between castor bean and arbuscular mycorrhizal (AM) fungi. The phenolic content in stressed roots was significantly lower compared to wet conditions with a negative correlation between AM fungal colonization and root phenolic content
... Counting phytoplankton to the species or genus level has high costs as it is time consuming and requires highly trained technicians (Edler and Elbrächter, 2010). The limited budget allocated to science and technology in Latin America likely prevent the incorporation of molecular techniques that have become increasingly popular to detect potential toxic cyanobacteria in monitoring programs (Moreira et al., 2014;Rosa et al., 2011;Salazar-Alcaraz et al., 2021;Salmaso et al., 2017). ...
Cyanobacterial blooms have serious adverse effects on human and environmental health. In Latin America, one of the main world's freshwater reserves, information on this phenomenon remains sparse. To assess the current situation, we gathered reports of cyanobacterial blooms and associated cyanotoxins in freshwater bodies from South America and the Caribbean (Latitude 22° N to 45° S) and compiled the regulation and monitoring procedures implemented in each country. As the operational definition of what is a cyanobacterial bloom remains controversial, we also analyzed the criteria used to determine the phenomena in the region. From 2000 to 2019, blooms were reported in 295 water bodies distributed in 14 countries, including shallow and deep lakes, reservoirs, and rivers. Cyanotoxins were found in nine countries and high concentrations of microcystins were reported in all types of water bodies. Blooms were defined according to different, and sometimes arbitrary criteria including qualitative (changes in water color, scum presence), quantitative (abundance), or both. We found 13 different cell abundance thresholds defining bloom events, from 2 × 103 to 1 × 107 cells mL−1. The use of different criteria hampers the estimation of bloom occurrence, and consequently the associated risks and economic impacts. The large differences between countries in terms of number of studies, monitoring efforts, public access to the data and regulations regarding cyanobacteria and cyanotoxins highlights the need to rethink cyanobacterial bloom monitoring, seeking common criteria. General policies leading to solid frameworks based on defined criteria are needed to improve the assessment of cyanobacterial blooms in Latin America. This review represents a starting point toward common approaches for cyanobacterial monitoring and risk assessment, needed to improve regional environmental policies.
... Se prepararon los siguientes medios de cultivos: Chlorophyta: BG11 (Allen, 1968;Andrade et al., 2009) y Cianobacteria: Zarrouk (1966) (Pineda et al., 2011). Los componentes de cada medio se presentan en el Anexo 1 y 2. Posteriormente, se ajustó BG11a pH 7.8 ‰ y Zarrouk a pH 9.9 ‰. ...
The industrial interest in phytoplanktonic microorganisms such as microalgae and cyanobacteria are due to the high biotechnological potential they have in synthesizing bioactive compounds, ease of adapting to different sources of nutrients, extreme survival conditions and the bioabsorbent capacity of heavy metals mainly in wastewater. The objective of this study was to isolate and characterize phenotypically and genotypically phytoplanktonic microorganisms from stabilization pools in Engabao, Guayas-Ecuador, using different culture media and molecular markers ITS, ITS2, LSU and 16S RNAr. The results of the cell culture during seven days showed the Chls1 microalgae with BG11 pH 7.8 medium reached 25.4 x 106 cells mL-1, the cyanobacterium Syn1 was 340.6 x 106 cells mL-1, while for Nod1 chlorophyll analysis was used to reach on the seventh day 6.33 μg mL-1. Molecular studies were carried out using a modified gDNA extraction and purification protocol, obtaining optimal DNA concentration and quality for the three strains: Chls1 146 ng μL-1, Syn1 326 ng μL-1 and Nod1 158.8 ng μL-1. The use of polyphasic analysis based on morphological characters and gDNA of the strains, was amplified by PCR and sequenced; the strains identified phenotypically and genotypically by the primers ITS2, ITS and LSU correspond to the microalgae Chlorella sorokiniana with 99.48%, 99.07% and 99.77% identity similarity, while the cyanobacterial isolates Synechococcus sp. and Nodosilinea sp. with the analysis of the 16S RNAr region, it presents identity percentages of 97.47% and 99.83%; despite not using specific primers for these microorganisms, the use of several regions increases the reliability of taxonomic and phylogenetic identification; these species have a high biotechnological potential and a great economic impact.
... En México existen lagos urbanos con florecimientos frecuentes o permanentes, que están destinados a actividades deportivas y/o recreativas y que representan por lo tanto un riesgo latente para la biota residente y circundante. Se sabe que concentraciones altas de nutrientes favorecen el crecimiento acelerado del fitoplancton, siendo las cianobacterias los microorganismos dominantes en los cuerpos de agua eutróficas ocasionando la formación de cúmulos de biomasa, conocidos como blooms o florecimientos (Pineda et al., 2011). ...
p>El lago urbano estudiado, está situado al oeste de la Ciudad de México. La problemática de la eutrofización del lago se agudizó y el lago presentó algunas características del estado de hipertrofía por su someridad y circulación limitada, falta de estratificación, regímenes de nutrientes y oxígeno no balanceado y productividad muy alta, tomando como parámetro concentraciones de Clorofila a . Por tal motivo, desde el 2015 hasta la actualidad, el IMTA viene monitoreando la calidad del agua de este lago y en marzo de 2016, se adaptó una planta piloto de coagulación-floculación alimentada con agua del mismo lago para el retiro de algas y el agua tratada se regresa a los lagos. Los resultados obtenidos son: el pH y la temperatura no tuvieron grandes variaciones durante éste periodo. Donde se observó una fuerte mejoría es en la calidad del agua de los lagos, fue en el aumento de la transparencia, y una considerable reducción en turbiedad y clorofila a . Reduciéndose también las concentraciones de nitrógeno total y fósforo total. Por último, la evaluación del aluminio, por su presencia en el coagulante, y el retorno del agua a los lagos, después del proceso de coagulación-floculación, no los afecta. Las condiciones de hipertrofia se mantienen, pero con el tratamiento de las aguas se proporcionó un mejor aspecto a los lagos y se controló las floraciones de Microcystis spp . y el mal olor.
Por tal motivo, desde el 2015 hasta la actualidad, el IMTA viene monitoreando la calidad del agua de este lago y en marzo de 2016, se adaptó una planta piloto de coagulación-floculación alimentada con agua del mismo lago para el retiro de algas y el agua tratada se regresa a los lagos.
Los resultados obtenidos son: el pH y la temperatura no tuvieron grandes variaciones durante éste periodo. Donde se observó una fuerte mejoría es en la calidad del agua de los lagos, fue en el aumento de la transparencia, y una considerable reducción en turbiedad y clorofila a . Reduciéndose también las concentraciones de nitrógeno total y fósforo total. Por último, la evaluación del aluminio, por su presencia en el coagulante, y el retorno del agua a los lagos, después del proceso de coagulación-floculación, no los afecta.
Las condiciones de hipertrofia se mantienen, pero con el tratamiento de las aguas se proporcionó un mejor aspecto a los lagos y se controló las floraciones de Microcystis spp . y el mal olor.
... Molecular studies are still scarce in all countries reviewed herein though growing in the last years with the exception of Peru where they are absent as far as we know. These genetic works have confirmed species adscription unequivocally in some of the waterbodies by sequencing of phylogenetic markers (i.e., genes 16S RNA, cpcBA-IGS, rpoC1 or nifH among others) in isolated cultures from Spain (Casero et al., 2014;Cirés et al., 2014b;Lezcano et al., 2019), andMexico (Arzate-Cárdenas et al., 2010;Lind et al., 2016;Pineda-Mendoza et al., 2012;Pineda-Mendoza et al., 2011); as well as in mixed field populations in Chile (Nimptsch et al., 2016), Colombia (Hurtado-Alarcón and Polanía-Vorenberg, 2014), Mexico (Vasconcelos et al., 2010), and Spain (Barón-Sola et al., 2012;Ouahid et al., 2005). Interestingly, many of these studies also confirmed by PCR the presence and identity of genes involved in the biosynthesis of MCs (mcy genes) in Chile (Nimptsch et al., 2016), Colombia (Hurtado-Alarcón and Polanía-Vorenberg, 2014), and Mexico (Arzate-Cárdenas et al., 2010;Pineda-Mendoza et al., 2012;Vasconcelos et al., 2010) and of MCs, CYN (cyr genes) and STXs (sxt genes) in Spain (Barón-Sola et al., 2012;Casero et al., 2014;Cirés et al., 2014b) offering a rapid screening method to qualitatively trace toxic populations. ...
The increasing occurrence of cyanobacterial blooms worldwide represents an important threat for both the environment and public health. In this context, the development of risk analysis and management tools as well as sustainable and cost-effective treatment processes is essential. The research project TALGENTOX, funded by the Ibero-American Science and Technology Program for Development (CYTED-2019), aims to address this ambitious challenge in countries with different environmental and social conditions within the Ibero-American context. It is based on a multidisciplinary approach that combines ecology, water management and technology fields, and includes research groups from Chile, Colombia, Mexico, Peru and Spain. In this review, the occurrence of toxic cyanobacteria and cyanotoxins in freshwaters from these countries are summarized. The presence of cyanotoxins has been confirmed in all countries but the information is still scarce and further monitoring is required. In this regard, remote sensing or metagenomics are good alternatives at reasonable cost. The risk management of freshwaters from those countries considering the most frequent uses (consumption and recreation) has been also evaluated. Only Spain and Peru include cyanotoxins in its drinking water legislation (only MC-LR) and thus, there is a need for regulatory improvements. The development of preventive strategies like diminishing nutrient loads to aquatic systems is also required. In the same line, corrective
... Among aquatic microorganisms, cyanobacteria are prokaryotic cells with a wide morphological diversity including unicellular, colonial, and filamentous forms. They are distributed in terrestrial, freshwater, and saltwater, and even in extreme environments [1]. Nitrogen and phosphorus are essential nutrients for aquatic microorganism growth. ...
In prokaryotes, the autotrophic population within the bacteria kingdom carries great biotechnological potential. For taxonomic analysis of species, the sequence encoding the small subunit ribosomal RNA gene (16S rRNA) is currently the most reliable technique for phylogenetic bacterial classification. Sequencing can define the population diversity and ecological importance within a habitat. The study of autotrophic bacteria population dynamics in a photobioreactor using a carbon-and nitrogen-free culture medium could provide information about succession dynamics under controlled conditions. The aim of this study was to identify the bacterial populations present in a photobioreactor by 16S rRNA amplicon sequence comparison from samples taken at 0, 7, and 14 fermentation days. Most of the identified clones were described to the level of genera from the following phyla: Proteobacteria (40%), Armatimonadetes (35%), Firmicutes (17.5%), Actinobacteria (2.5%), and Cyanobacteria/Chloroplast (2.5%). Bacterial strains from such phyla have been reported in aquatic environments and were considered uncultivable; however, sequences indicated the presence of the clones A-123 (ID: HQ860522.1) from time 0, and YHS25 (GU305825.1) after 7 days of culture. Our findings demonstrated the major presence of autotrophic bacterial populations of the Proteobacteria and Armatimonadetes phyla, which grew under artificial culture conditions in a photobioreactor.
Este trabajo presenta una aproximación al conocimiento generado en relación con el género Guadua a partir del contexto histórico e incluyendo el estado de arte que demarca las oportunidades de investigación. En consecuencia, se inicia con el análisis de diversas fuentes de la historia ambiental para el abordaje de los usos ancestrales de la guadua, que progresivamente dejan entrever la inserción de este bien natural en los escenarios sociales, políticos y culturales de las comunidades. Se continúa con la revisión de referentes bibliográficos de los últimos diez años, identificando enfoques disciplinares sobresalientes en el estudio de la guadua, así como amplia documentación de las comunidades del Centro y Occidente colombiano con Guadua angustifolia Kunth como protagonista, lo que establece una oportunidad para la generación de conocimiento en la Región Caribe colombiana. Por ende, este trabajo constituye la apertura hacia un tema de investigación conducente a indagar el papel que desempeña en la Región Caribe colombiana, la especie de guadua nativa identificada como Guadua amplexifolia J. Presl desde la dimensión sociocultural de las comunidades en esa región del país.
Artificial urban lakes commonly have physicochemical conditions that contribute to rapid anthropogenic eutrophication and development of cyanobacterial blooms. Microcystis is the dominat genus in the most freshwater bodies and is one of the main producter of microcystins. Using 454-pyrosequencing we characterized the bacterial community, with special emphasis on Microcystis, in three recreational urban lakes from Mexico City in both wet and dry seasons. We also evaluated some physicochemical parameters that might influence the presence of Microcystis blooms, and we associated the relative abundance of heterotrophic and autotrophic bacterial communities with their possible metabolic capacities. A total of 14 phyla, 18 classes, 39 orders, 53 families and 48 bacterial genera were identified in both seasons in the three urban lakes. Cyanobacteria had the highest relative abundance followed by Proteobacteria and Actinobacteria. Microcystis was the dominant taxon followed by Arthrospira, Planktothrix and Synechococcus. We also found heterotrophic bacteria associated with the blooms, such as Rhodobacter, Pseudomonas, Sphingomonas and, Porphyrobacter. The highest richness, diversity and dominance were registered in the bacterial community of the Virgilio Uribe Olympic Rowing-Canoeing Track in both seasons, and the lowest values were found in the Chapultepec Lake. The canonical correspondence analysis showed that dissolved oxygen and NO3–N concentrations might explain the presence of Microcystis blooms. The metabolic prediction indicated that these communities are involved in photosynthesis, oxidative phosphorylation, methane metabolism, carbon fixation, and nitrogen and sulfur metabolism. The lakes studied had a high prevalence of Microcystis, but average values of microcystins did not exceed the maximum permissible level established by the United States Environmental Protection Agency for recreational and cultural activities. The presence of cyanobacteria and microcystins at low to moderate concentrations in the three lakes could result in ecosystem disruption and increase animal and human health risks.
The following investigation took place in the lake Parque Alameda de la Solidaridad in Cuernavaca Morelos. Samples at three sites in the lake were taken to determine the variation and abundance of phytoplankton during the four seasons the year. Temperature, transparency, oxygen, pH and nutrients (nitrates, amonium and phosphorous) were determined. Thirty species were recognized, the Chlorophyceae were the most abundant and with the higher number of species. The higher abundances of all the period were in spring and winter. Oocystis solitaria and Leptolyngbya sp. were the predominant species with abundances higher than (> 1x10(6) org/ml). The variation of phycoflora in the lake, showed a temporal succession in frequent, seasonal, occasional and rare species. The phytoplankton species richness and abundance, as well nutrients concentrations showed the lake as a hypertrophic system water body.
Five strains that were morphologically identified as Planktolyngbya circumcreta were isolated from three geographical regions in China. The strains were phylogenetically and taxonomically characterised based on a polyphasic approach that merged morphological and genetic characteristics, as well as ultrastructure. The ultrastructure of these strains showed a similar arrangement of thylakoids, with four to eight parallel layers that were parietally distributed, suggesting that they belonged to the order Synechococcales, family Leptolyngbyaceae. As inferred from 16S rRNA gene sequence analysis, these pure cultures share ≥ 98.5% similarity and were grouped into a unique and robust cluster with high bootstrap values. This unique cluster was separated from a clade composed of two strains of Planktolyngbya limnetica, which belonged to the type species of the genus Planktolyngbya. Thus, we transferred these five P. circumcreta strains to the new genus Limnolyngbya. The species Limnolyngbya circumcreta nov. comb. was designated as the type of the genus.
The following investigation took place in the lake Parque Alameda de la Solidaridad in Cuernavaca Morelos. Samples at three sites in the lake were taken to determine the variation and abundance of phytoplankton during the four seasons the year. Temperature, transparency, oxygen, pH and nutrients (nitrates, amonium and phosphorous) were determined. Thirty species were recognized, the Chlorophyceae were the most abundant and with the higher number of species. The higher abundances of all the period were in spring and winter. Oocystis solitaria and Leptolyngbya sp. were the predominant species with abundances higher than (>1×10⁶ org/ml). The variation of phycoflora in the lake, showed a temporal succession in frequent, seasonal, occasional and rare species. The phytoplankton species richness and abundance, as well nutrients concentrations showed the lake as a hypertrophic system water body.
The cyanobacteria are photosynthetic prokaryotes possessing the ability to synthesize chlorophyll a and at least one phycobilin pigment; typically water acts as the electron donor during photosynthesis, leading to the release of oxygen. They are by far the largest group of photosynthetic prokaryotes, as judged by their widespread occurrence, frequent abundance, and morphological diversity. Not only are they represented at the present day in most types of illuminated environment, except for those at lower pH values, but they have one of the longest geological records (Schopf and Walter, 1982). Much of the earth’s original atmospheric oxygen was probably formed by organisms quite similar to modern cyanobacteria (Knoll, 1985) and they are still responsible for a considerable proportion of photosynthetic oxygen evolution in the oceans.
Palabras clave: Cianobacterias, toxicidad, aguas continentales, Chile Cyanobacteria are phototropic and oxygenic microorganisms that are widely distributed and reported to be present in freshwater or seawater from the five continents. They can eventually develop into large masses called «blooms» which are usually generated by anthropogenic effects. Several species of cyanobacteria can produce potent neurotoxins and hepatotoxins, which are difficult to eliminate from water. The intoxica-tion with some of these toxins can become severe causing death in animals and man and it is suspected that ingestion of even microdoses of them can induce tumor appearance. The distribution of cyanobacteria in continental waters of Chile has been recorded from uppermost north of Chile up to the Antarctic territory. The presence of toxic species as well as the development of blooms have been detected in different regions.The mycrocystin cyanotoxin has been assessed in different masses of water by means of high pressure liquid chromatography (HPLC) together with mass spectrometry (MALDI-TOF) (PSD). Finally it is recommended to make analysis in order to establish controls that avoid risk of exposure to these cyanotoxins.
Cyanobacteria are known to produce secondary metabolites which are toxic to mammals, commonly known as "toxins". These have been described as having neurotoxic, hepatotoxic and dermatotoxic effects, being a hazard also to humans. Cyanobacteria blooms may represent a hazard to aquatic organisms, when cells collapse, releasing cyanotoxins to the water. Toxins, though, have a dramatic influence on many groups of other arganisms. Zooplankton species are affected by cyanotoxins Duphniu is used as a test organism for cyanobacteria toxicity evaluation. Rotifers and small cladocerans seem to be less sensitive to cyanobacteria toxicity. This partly explains the dominance of small zooplankton groups during cyanobacteria blooms. Molluscs such as mussels are also little affected by cyanobacteria toxins, being able to transfer them along food chains up to humans. Mussels purify microcystins from their body slowly, making them good toxin vectors. Larger crustaceans such as crayfish may be also considered as toxin vectors, having low toxin sensitivity even during early larval phases. The response of fish to toxins is quite diverse and is related to food habit. Carp seem to accumulate more toxins than mullet or barb and they are not much affected by oral toxicity. Many of these animals may act as vectors for toxins better than other more sensitive organisms, which may be badly affected by the toxins.
The production of potent toxins by bloom-, scum- and mat-forming cyanobacteria, in fresh-, brackish and marine waters, appears to be a global phenomenon. Cyanobacterial toxins can also be produced by cyanobacteria from terrestrial sources. The range and number of known cyanobacterial toxins are increasing apace as associated poisoning incidents are investigated, and increasingly powerful analytical methods are applied to complement toxicity-based studies on both natural samples and laboratory isolates of cyanobacteria. Water quality management to reduce toxic cyanobacterial mass developments, and schemes to mitigate the potential effects of cyanobacterial toxins, require an understanding of the occurrence and properties of the toxins and of the exposure routes via which the toxins present risks to health. Here, we review advances in the recognition of cyanobacterial toxins and their toxicity, and of the exposure routes with reference to human health, namely via skin contact, inhalation, haemodialysis and ingestion (the oral route).
An approximately 1400-bp region of the 16S rRNA gene was sequenced for 10 polar or near-polar strains putatively placed in the Oscillatorialean genera Oscillatoria, Phormidium, and Lyngbya obtained from the University of Toronto Culture Collection to assess phylogenetic relationships. The strains were also examined for thylakoid structure and cell division type with TEM as well as traditional morphology with LM. Phylogenetic trees constructed using parsimony, distance, and maximum likelihood methods were similar in topology. If the original epithets applied to the sequenced strains (both polar and those from GenBank) were used, it was clear that taxa were not monophyletic. However, using the revised taxonomic system of Anagnostidis and Komárek, we were able to reassign these strains to their current correct taxa (species, genus, and family). When these assignments were made, it was determined that the molecular sequence data analyses were congruent with morphology and ultrastructure. Nine of the polar strains were found to be new species, and eight were described as such: Arthronema gygaxiana Casamatta et Johansen sp. nov., Pseudanabaena tremula Johansen et Casamatta sp. nov., Leptolyngbya angustata Casamatta et Johansen sp. nov., Phormidium lumbricale Johansen et Casamatta sp. nov., Microcoleus glaciei Johansen et Casamatta sp. nov., Microcoleus rushforthii Johansen et Casamatta sp. nov., Microcoleus antarcticus Casamatta et Johansen sp. nov., Microcoleus acremannii Casamatta et Johansen sp. nov. Some genera (Leptolyngbya and Microcoleus) were clearly not monophyletic and require future revision.
Microcystis aeruginosa, a cosmopolitan form, is a colonial cyanobacterium, which is also common in many freshwater bodies in Mexico. In eutrophic
water bodies cyanobacteria are often the main phytoplankton that co-exist with cladocerans. We evaluated the effect of mixed
diets, comprising 0, 25, 50, 75, and 100% on dry weight basis of M. aeruginosa, and the rest of one of two green algal species (Chlorella vulgaris and Scenedesmus acutus), on the population growth of the cladocerans Ceriodaphnia dubia and Moina macrocopa. Regardless of the share of M. aeruginosa in the mixed diet, C. dubia fed Chlorella had a longer initial lag phase. However, in mixed diet with S. acutus, the lag phase of C. dubia increased with increasing proportion of M. aeruginosa. When raised on 100% M. aeruginosa, the population growth of C. dubia was lowered compared with 100% S. acutus or 100% C. vulgaris. Increased proportion of M. aeruginosa in the mixed diet also resulted in decreased abundance of M. macrocopa. Irrespective of diet type, M. macrocopa had a shorter lag phase than C. dubia. Depending on the diet type, the rate of population increase (r) of C. dubia varied from 0.07 to 0.26d−1 while that of M. macrocopa was higher (0.14–0.61d−1). For both cladoceran species, the lower r values were obtained when fed Microcystis. Our study showed that the strain of M. aeruginosa was not highly toxic to cause total elimination of either C. dubia or M. macrocopa. Addition of a green algal component to the diet improved the population growth rates of both cladoceran species.
Dense surface blooms of toxic cyanobacteria in eutrophic lakes may lead to mass mortalities of fish and birds, and provide a serious health threat for cattle, pets, and humans. It has been argued that global warming may increase the incidence of harmful algal blooms. Here, we report on a lake experiment where intermittent artificial mixing failed to control blooms of the harmful cyanobacterium Microcystis during the summer of 2003, one of the hottest summers ever recorded in Europe. To understand this failure, we develop a coupled biological-physical model investigating how competition for light between buoyant cyanobacteria, diatoms, and green algae in eutrophic lakes is affected by the meteorological conditions of this extreme summer heatwave. The model consists of a phytoplankton competition model coupled to a one-dimensional hydrodynamic model, driven by meteorological data. The model predicts that high temperatures favour cyanobacteria directly, through increased growth rates. Moreover, high temperatures also increase the stability of the water column, thereby reducing vertical turbulent mixing, which shifts the competitive balance in favour of buoyant cyanobacteria. Through these direct and indirect temperature effects, in combination with reduced wind speed and reduced cloudiness, summer heatwaves boost the development of harmful cyanobacterial blooms. These findings warn that climate change is likely to yield an increased threat of harmful cyanobacteria in eutrophic freshwater ecosystems.
The Holliday junction is a central intermediate in genetic recombination. This four-stranded DNA structure is capable of spontaneous
branch migration, and is lost during standard DNA extraction protocols. In order to isolate and characterize recombination
intermediates that contain Holliday junctions, we have developed a rapid protocol that restrains branch migration of four-way
DNA junctions. The cationic detergent hex-adecyltrimethylammonium bromide is used to lyse cells and precipitate DNA. Manipulations
are performed in the presence of the cations hexamine cobalt(III) or magnesium, which stabilize Holliday junctions in a stacked-X
configuration that branch migrates very slowly. This protocol was evaluated using a sensitive assay for spontaneous branch
migration, and was shown to preserve both artificial Holliday junctions and meiotic recombination intermediates containing
four-way junctions.
Microcystis is a bloom-forming, common cyanobacterium in urban lakes of Mexico City. To assess the presence of potentially cyanotoxin-producing Microcystis, molecular techniques were applied and acute toxicity bioassays were performed with Daphnia magna neonates exposed to cyanobacterial crude extracts. Toxigenic potential of isolated strains was inferred by amplifying the mcyA-Cd genes and their identity as Microcystis was confirmed through the 16S rDNA and phycocyanin operon amplification. Microcystins synthesized under culture conditions were quantified through ELISA. The acute toxicity bioassays revealed that mortality was independent from the cyanotoxin concentration in some strains; this suggests the presence of other metabolites (different from microcystins) that also exerted an important biological effect. Isolated strains had the mcyA-Cd gene and most of them produced variable amounts of microcystins in the culture conditions used, confirming their toxigenic potential. Results warn about possible toxic effect risks for aquatic biota, neighboring areas, visitors and users of these sites, due to the constant presence of these blooms in the studied water bodies.
The increasing eutrophication of the fresh water reservoirs has lead to the frequent occurrence of uncontrolled proliferations of phytoplankton, mainly of cyanobacteria, which can have toxic character. One of the most virulent strains is the genera Microcystis which produces the most prevalent cyanotoxin, the microcystin. The microcystins pose a serious health and environmental risk, and the Spanish current legislation already controls the level of the toxin in potable waters. There are two main open questions about microcystins that have been long discussed and still remain unresolved. Firstly, what the factors that trigger microcystin synthesis are, and secondly, what the physiological role of these molecules could be in the cyanobacteria, since they are mainly found as endotoxins. La creciente eutrofización de los acuíferos ha provocado que frecuentemente se produzcan proliferaciones incontroladas de fitoplancton, principalmente de cianobacterias, que pueden tener carácter tóxico. Una de las cepas más virulentas es del género Microcystis y produce la cianotoxina más ubicua, la microcistina. Las microcistinas producen graves problemas sanitarios y medioambientales, y la legislación Española contempla ya su control en aguas emergentes de depuradoras. Hay dos grandes problemas planteados en torno a la microcistina. En primer lugar se desconoce qu¿e factores son los que desencadenan su síntesis, y en segundo lugar, qué papel fisiológico juegan en la cianobacteria, ya que se trata mayoritariamente de una endotoxina.
Con base en los resultados de 16 muestreos mensuales de los nutrimentos (fósforo y nitrógeno) y de la clorofila a en el Lago Viejo de Chapultepec en la Ciudad de México, que es un sistema acuático somero recreativo y con algunas caracteristicas de hipertrofia, se evaluó el efecto ejercidc por el dragado de una zona del Lago, medida llevada a cabo para restaurarlo. No obstante que los valores de nitratos, ortofosfatos, fósforo total y clorofüa a aumentaron (40, 2, 1.5 y 2 veces, respectivamente) después del dragado con relación a los meses inmediatos anteriores, se consideró que los valores de los parámetros anteriormente citados quedaron comprendidos en la variación temporal normal presente antes del mismo. Se discute la pertinencia del dragado como medida correctiva de los efectos negativos provocados por las condiciones eutróficas y se concluye que se requieren monitoreos extensos para evaluar con eficiencia los resultados de las acciones emprendidas.
In order to assess the potential of several molecular targets for the identification, typing and traceability of cyanobacteria in freshwater reservoirs, molecular techniques were applied to 118 cyanobacterial isolates mostly sourced from Portuguese freshwater reservoirs and representative of three orders of cyanobacteria: Chroococcales (54), Oscillatoriales (15) and Nostocales (49). The isolates were previously identified by morphological methods and subsequently characterized by composite hierarchical cluster analysis of STRR and LTRR (short and long tandemly repeated repetitive sequences) PCR fingerprinting profiles. Representative isolates were selected from each cluster and their molecular identification, at the species level, was obtained or confirmed by phylogenetic positioning using 16S rRNA gene and rpoC1 phylogenies. A highly congruent association was observed between STTR- and LTRR-based clusters and taxonomic affiliation, revealing the usefulness of such PCR fingerprinting profiles for the identification of cyanobacteria. Composite analysis of hierarchical clustering of M13 and ERIC PCR fingerprints also appeared suitable for strain typing and traceability within a reservoir, indicating its potential for use in cyanobacterial monitoring, as a quality management control. Based on Simpson (D) and Shannon-Wiener (J') indices a high diversity was observed within all species, with Planktothrix agardhii showing the lowest diversity values (D=0.83; J'=0.88) and Aphanizomenon flos-aquae the highest ones (D=J'=0.99). A diagnostic key based on 16S-ARDRA, ITS amplification and ITS-ARDRA for identification purposes is also presented.
Waterbloom samples of the colonial cyanobacterium Microcystis aeruginosa, collected in fish ponds at the Hydrobiological Institute, Wuhan, People's Republic of China, were hepatotoxic to mice. Lyophilized cells had an LD50 (i.p. mouse; 40 mg/kg) and signs of poisoning similar to that reported for other cyanobacterial hepatotoxic peptides. Two toxins, with an LD50 (i.p. mouse) of 40 and 150 micrograms/kg, were isolated using gel filtration and high performance liquid chromatography. The amino acid composition and mol. wt (994) of the 40 micrograms/kg toxin was the same as that for microcystin-LR, while the 150 micrograms/kg toxin had an amino acid composition and mol. wt (1048) different from any of the reported cyanobacteria heptapeptide toxins reported to date.
A major problem in development of a polyphasic taxonomy is that the identification of oxyphotobacterial strains (cyanobacteria and prochlorophytes) in culture collections may be incorrect. We have therefore developed a diagnostic system using the DNA sequence polymorphism in the 16S rRNA regions V6 to V8 for individual strain characterization and identification. PCR primers amplifying V6 to V8 from oxyphotobacteria in unialgal cultures were constructed. Direct solid-phase or cyclic sequencing was used to determine the sequences from the amplified DNA. This survey includes 10 strains of Nostoc/Anabaena/Aphanizomenon (Nostoc category), 5 strains of Microcystis (Microcystis category), and 4 strains of Planktothrix (Planktothrix category). Fifteen additional strains of cyanobacteria and two strains of prochlorophytes were included such that the major phyletic groups were represented. One of the strains, Phormidium sp. NIVA-CYA 203, contained an 11-nucleotide insertion with no homology to other known 16S rRNA sequences. Based on parsimony and neighbor-joining trees, the phyletic relationships of the strains were investigated. Thirteen major branches were found, with Pseudanabaena limnetica NIVA-CYA 276/6 as the most divergent strain. The strain categories Nostoc, Planktothrix, and Microcystis were all monophyletic. The sequence polymorphism within Nostoc was higher than that in Planktothrix and Microcystis. Based on the sequence and phyletic information, group-specific PCR primers for the categories Nostoc, Planktothrix, and Microcystis were constructed. For the strains included in this work, the amplifications were specific for the relevant groups. By combination of magnetic solid-phase DNA isolation and group-specific PCR amplifications, an accurate method for characterization, classification and identification of oxyphotobacterial clone cultures has been developed.
We developed and tested a set of oligonucleotide primers for the specific amplification of 16S rRNA gene segments from cyanobacteria and plastids by PCR. PCR products were recovered from all cultures of cyanobacteria and diatoms that were checked but not from other bacteria and archaea. Gene segments selectively retrieved from cyanobacteria and diatoms in unialgal but nonaxenic cultures and from cyanobionts in lichens could be directly sequenced. In the context of growing sequence databases, this procedure allows rapid and phylogenetically meaningful identification without pure cultures or molecular cloning. We demonstrate the use of this specific PCR in combination with denaturing gradient gel electrophoresis to probe the diversity of oxygenic phototrophic microorganisms in cultures, lichens, and complex microbial communities.
CLUSTAL X is a new windows interface for the widely-used progressive multiple sequence alignment program CLUSTAL W. The new
system is easy to use, providing an integrated system for performing multiple sequence and profile alignments and analysing
the results. CLUSTAL X displays the sequence alignment in a window on the screen. A versatile sequence colouring scheme allows
the user to highlight conserved features in the alignment. Pull-down menus provide all the options required for traditional
multiple sequence and profile alignment. New features include: the ability to cut-and-paste sequences to change the order
of the alignment, selection of a subset of the sequences to be realigned, and selection of a sub-range of the alignment to
be realigned and inserted back into the original alignment. Alignment quality analysis can be performed and low-scoring segments
or exceptional residues can be highlighted. Quality analysis and realignment of selected residue ranges provide the user with
a powerful tool to improve and refine difficult alignments and to trap errors in input sequences. CLUSTAL X has been compiled
on SUN Solaris, IRIX5.3 on Silicon Graphics, Digital UNIX on DECstations, Microsoft Windows (32 bit) for PCs, Linux ELF for
x86 PCs, and Macintosh PowerMac.
Nonribosomal peptide synthesis is achieved in prokaryotes and lower eukaryotes by the thiotemplate function of large, modular enzyme complexes known collectively as peptide synthetases. These and other multifunctional enzyme complexes, such as polyketide synthases, are of interest due to their use in unnatural-product or combinatorial biosynthesis (R. McDaniel, S. Ebert-Khosla, D. A. Hopwood, and C. Khosla, Science 262:1546-1557, 1993; T. Stachelhaus, A. Schneider, and M. A. Marahiel, Science 269:69-72, 1995). Most nonribosomal peptides from microorganisms are classified as secondary metabolites; that is, they rarely have a role in primary metabolism, growth, or reproduction but have evolved to somehow benefit the producing organisms. Cyanobacteria produce a myriad array of secondary metabolites, including alkaloids, polyketides, and nonribosomal peptides, some of which are potent toxins. This paper addresses the molecular genetic basis of nonribosomal peptide synthesis in diverse species of cyanobacteria. Amplification of peptide synthetase genes was achieved by use of degenerate primers directed to conserved functional motifs of these modular enzyme complexes. Specific detection of the gene cluster encoding the biosynthetic pathway of the cyanobacterial toxin microcystin was shown for both cultured and uncultured samples. Blot hybridizations, DNA amplifications, sequencing, and evolutionary analysis revealed a broad distribution of peptide synthetase gene orthologues in cyanobacteria. The results demonstrate a molecular approach to assessing preexpression microbial functional diversity in uncultured cyanobacteria. The nonribosomal peptide biosynthetic pathways detected may lead to the discovery and engineering of novel antibiotics, immunosuppressants, or antiviral agents.
Toxic and non-toxic cyanobacterial strains from Anabaena, Aphanizomenon, Calothrix, Cylindrospermum, Nostoc, Microcystis, Planktothrix (Oscillatoria agardhii), Oscillatoria and Synechococcus genera were examined by RFLP of PCR-amplified 16S rRNA genes and 16S rRNA gene sequencing. With both methods, high 16S rRNA gene similarity was found among planktic, anatoxin-a-producing Anabaena and non-toxic Aphanizomenon, microcystin-producing and non-toxic Microcystis, and microcystin-producing and non-toxic Planktothrix strains of different geographical origins. The respective sequence similarities were 99.9-100%, 94.2-99.9% and 99.3-100%. Thus the morphological characteristics (e.g. Anabaena and Aphanizomenon), the physiological (toxicity) characteristics or the geographical origins did not reflect the level of 16S rRNA gene relatedness of the closely related strains studied. In addition, cyanobacterial strains were fingerprinted with repetitive extragenic palindromic (REP)- and enterobacterial repetitive intergenic consensus (ERIC)-PCR. All the strains except two identical pairs of Microcystis strains had different band profiles. The overall grouping of the trees from the 16S rRNA gene and the REP- and ERIC-PCR analyses was similar. Based on the 16S rRNA gene sequence analysis, four major clades were formed. (i) The clade containing filamentous heterocystous cyanobacteria was divided into three discrete groups of Anabaena/Aphanizomenon, Anabaena/Cylindrospermum/ Nodularia/Nostoc and Calothrix strains. The three other clades contained (ii) filamentous non-heterocystous Planktothrix, (iii) unicellular non-heterocystous Microcystis and (iv) Synechococcus strains.
Nodularia, a member of the order Nostocales, is a bloom-forming filamentous cyanobacterium that possesses the ability to form toxic blooms. The toxin produced by Nodularia, nodularin, is a hepatotoxin, similar in structure to the heptapeptide toxin microcystin. Twenty-one strains of Nodularia, representing the species Nodularia spumigena, Nodularia harveyana and Nodularia sphaerocarpa, were analysed for toxin production by protein phosphatase inhibition assay and sequenced over the 16S rDNA region. Phylogenetic analysis of Nodularia 16S rDNA sequences found that Nodularia clustered into two main groups. An N. spumigena cluster was distinct from the benthic species N. harveyana and N. sphaerocarpa. There was no distinction between strains isolated from globally diverse locations. Nodularin-producing species were restricted to the single, evolutionally distinct cluster of N. spumigena. This observation has enabled the design of a specific 16S rRNA PCR for the rapid detection of nodularin-producing strains. Alignment of 16S rDNA sequences from toxic and non-toxic Nodularia with other members of the cyanobacteria allowed the design of both Nodularia generic and toxic N. spumigena-specific primers.
The rapid increase in the amount of protein and DNA sequence information available has become almost overwhelming to researchers. So much information is now accessible that high-quality, functional gene analysis and categorization has become a major goal for many laboratories. To aid in this categorization, there is a need for non-commercial software that is able to both align sequences and also calculate pairwise levels of similarity/identity.
We have developed MatGAT (Matrix Global Alignment Tool), a simple, easy to use computer application that generates similarity/identity matrices for DNA or protein sequences without needing pre-alignment of the data.
The advantages of this program over other software are that it is open-source freeware, can analyze a large number of sequences simultaneously, can visualize both sequence alignment and similarity/identity values concurrently, employs global alignment in calculations, and has been formatted to run under both the Unix and the Microsoft Windows Operating Systems. We are presently completing the Macintosh-based version of the program.
Although diverse natural products have been isolated from the benthic, filamentous cyanobacterium Lyngbya majuscula, it is unclear whether this chemical variation can be used to establish taxonomic relationships among disparate collections.
We compared morphological characteristics, secondary-metabolite compositions, and partial 16S ribosomal DNA (rDNA) sequences
among several collections of L. majuscula Gomont, Lyngbya spp., and Symploca spp. from Guam and the Republic of Palau. The morphological characteristics examined were cell length, cell width, and the
presence or absence of a calyptra. Secondary metabolites were analyzed by two-dimensional thin-layer chromatography. Each
collection possessed a distinct cellular morphology that readily distinguished Lyngbya spp. from Symploca spp. Each collection yielded a unique chemotype, but common chemical characteristics were shared among four collections of
L. majuscula. A phylogeny based on secondary-metabolite composition supported the reciprocal monophyly of Lyngbya and Symploca but yielded a basal polytomy for Lyngbya. Pairwise sequence divergence among species ranged from 10 to 14% across 605 bp of 16S rDNA, while collections of L. majuscula showed 0 to 1.3% divergence. Although the phylogeny of 16S rDNA sequences strongly supported the reciprocal monophyly of
Lyngbya and Symploca as well as the monophyly of Lyngbya bouillonii and L. majuscula, genetic divergence was not correlated with chemical and morphological differences. These data suggest that 16S rDNA sequence
analyses do not predict chemical variability among Lyngbya species. Other mechanisms, including higher rates of evolution for biosynthetic genes, horizontal gene transfer, and interactions
between different genotypes and environmental conditions, may play important roles in generating qualitative and quantitative
chemical variation within and among Lyngbya species.
The heterocytous cyanobacteria form a monophyletic group according to 16S rRNA gene sequence data. Within this group, phylogenetic and morphological studies have shown that genera such as Anabaena and Aphanizomenon are intermixed. Moreover, the phylogeny of the genus Trichormus, which was recently separated from Anabaena, has not been investigated. The aim was to study the taxonomy of the genera Anabaena, Aphanizomenon, Nostoc and Trichormus belonging to the family Nostocaceae (subsection IV.I) by morphological and phylogenetic analyses of 16S rRNA gene, rpoB and rbcLX sequences. New strains were isolated to avoid identification problems caused by morphological changes of strains during cultivation. Morphological and phylogenetic data showed that benthic and planktic Anabaena strains were intermixed. In addition, the present study confirmed that Anabaena and Aphanizomenon strains were not monophyletic, as previously demonstrated. The evolutionary distances between the strains indicated that the planktic Anabaena and Aphanizomenon strains as well as five benthic Anabaena strains in cluster 1 could be assigned to a single genus. On the basis of the 16S rRNA, rpoB and rbcLX gene sequences, the Anabaena/Aphanizomenon strains (cluster 1) were divided into nine supported subclusters which could also be separated morphologically, and which therefore might represent different species. Trichormus strains were morphologically and phylogenetically heterogeneous and did not form a monophyletic cluster. These Trichormus strains, which were representatives of three distinct species, might actually belong to three genera according to the evolutionary distances. Nostoc strains were also heterogeneous and seemed to form a monophyletic cluster, which may contain more than one genus. It was found that certain morphological features were stable and could be used to separate different phylogenetic clusters. For example, the width and the length of akinetes were useful features for classification of the Anabaena/Aphanizomenon strains in cluster 1. This morphological and phylogenetic study with fresh isolates showed that the current classification of these anabaenoid genera needs to be revised.
We have studied the diversity of pelagic cyanobacteria in Lake Loosdrecht, The Netherlands, through recovery and analysis of small subunit ribosomal RNA gene sequences from lake samples and cyanobacterial isolates. We used an adapted protocol for specific amplification of cyanobacterial rDNA for denaturing gradient gel electrophoresis (DGGE) analysis. This protocol enabled direct comparison of cyanobacterial community profiles with overall bacterial profiles. The theoretical amplification specificity of the primers was supported by sequence analysis of DNA from excised DGGE bands. Sequences recovered from these bands, in addition to sequences obtained by polymerase chain reaction (PCR) and cloning from lake DNA as well as from cyanobacterial isolates from the lake, revealed a diverse consortium of cyanobacteria, among which are representatives of the genera Aphanizomenon, Planktothrix, Microcystis and Synechococcus. One numerically important and persistent cyanobacterium in the lake, Prochlorothrix hollandica, appeared to co-occur with an unknown but related species. However, the lake is dominated by filamentous species that originally have been termed 'Oscillatoria limnetica-like'. We show that this is a group of several related cyanobacteria, co-occurring in the lake, which belong to the Limnothrix/Pseudanabaena group. The available variation among the coexisting strains of this group can explain the persistent dominance of the group under severe viral pressure.
Scoring the agar plate before incubation under unidirectional light led to a rapid separation of gliding filamentous cyanobacteria from their contaminating bacteria. Twenty strains were purified by the method. Additionally, 13 axenic cyanobacterial strains were isolated from pour plates made after treatment of cyanobacterial cultures in tryptone-yeast extract-glucose broth with cycloserine in darkness to select for obligate photoautotrophs.
The enrichment of water bodies with plant nutrients often results in mass growths of cyanobacteria which can produce a range of toxins. Cyanobacterial toxins have adverse effects on fish, birds, and mammals, and are being increasingly recognized as animal and human health hazards. The effects of cyanobacterial toxins on plants, particularly aquatic macrophytes, are little understood. The uptake of microcystin-LR (MC-LR) by Ceratophyllum demersum was detected with approximately 11.2% of the applied 14C-labelled microcystin being taken up after 7 days of exposure. The effects of various concentrations of this toxin on the detoxication enzyme glutathione S-transferase (microsomal and soluble forms) were investigated. Microsomal glutathione S-transferase activity was increased in C. demersum exposed to concentrations of MC-LR above 0.12 μg/L, but reduced at concentrations above 1.0 μg/L, whereas soluble glutathione S-transferase activity was increased by exposure to MC-LR concentrations above 0.5 μg/L. No reduction of soluble GST activity was observed. The activity of both microsomal and soluble GST systems, freshly prepared from a range of plants, was inhibited in the presence of 0.5 μg/L MC-LR. The results indicate a detoxication pathway for MC-LR in aquatic plants. ©1999 John Wiley & Sons, Inc. Environ Toxicol 14: 111–115, 1999
The role of external Ca2+ in the homeostasis of intracellular pH (pHi) of Anabaena sp. strain PCC7120 in response to a decrease in the external pH (pHex) has been studied in cell suspensions. Increase in cytoplasmic pH after acid shock is dependent on the presence of Ca2+ in the medium. The observed Ca2+-mediated alkalization of the cytoplasm depends on the extent of the shift in external pH. Acid pH shifts resulted in an increased permeability of the cytoplasmic membrane to protons, which could be reversed by increasing the concentration of Ca2+ in the medium. Thus, the ability of Ca2+ to increase cytoplasmic pH might be correlated with an inhibition of net proton uptake by increasing concentrations of external Ca2+ under these conditions. This combined response resulted in the generation and maintenance of a larger pH gradient (ΔpH) at acid external pH values. All Ca2+ channel blockers tested, such as verapamil and LaCl3, inhibited the observed Ca2+-mediated response. On the other hand, the Ca ionophore calcimycin (compound A23187) was agonistic, and stimulated both cytoplasmic alkalization and inhibition of net proton uptake. The protonophorous uncoupler carbonylcyanide m-chlorophenyl hydrazone, inhibited this Ca2+-mediated response, whereas monensin, an inhibitor of the Na+/H+ antiporter, had no significant effect. The results of the present study suggest that an influx of Ca2+ from the extracellular space is required for the regulation of cytoplasmic pH in Anabaena sp. strain PCC7120 exposed to low external pH values.
One of the most severe problems associated with eutrophication of urban freshwater ecosystems is the occurrence of increasingly frequent blooms of toxic cyanobacteria. Cyanotoxins might accumulate in the trophic web, producing diverse intoxication symptoms and chronic effects that are difficult to diagnose and prevent. High mortality of domestic animals and fish has been reported previously under these prevailing conditions. This study investigates the taxonomic composition of phytoplankton assemblages in Sheldon Lake during the summer of 2004, a year after the completion of a restoration project on the lake. The study analysed the physical and chemical changes caused by urban run-off and artificial mixing, as well as the usefulness of microcystin molecular markers derived from the mcy gene cluster for the detection of toxic cyanobacterial strains in environmental samples from Sheldon Lake. This study clearly demonstrates that the artificial mixing rate alone was insufficient to cause a transition to a well-mixed aquatic system, and that cyanobacteria remained dominant throughout the summer months. The presence of toxic cyanobacterial strains was confirmed with the use of molecular markers that detected the presence of the mcy gene cluster responsible for the production of toxin by Microcystis spp. This approach might have a great potential use in the routine analyses of urban aquatic ecosystems. It also might make toxicity monitoring more feasible, allowing for the early application of corrective actions, especially for cases such as Sheldon Lake, which is a public recreational focal point.
High molecular weight (HMW) DNA prepared from a toxic freshwater cyanobacterial bloom sample was used to construct a PCR-generated 75-clone, 16S rRNA gene library and a 2850-clone bacterial artificial chromosome (BAC) library. Phylogenetic analysis of the 16S rRNA gene library demonstrated that members of eight phyla of domain Bacteria, which included Cyanobacteria, Actinobacteria, Verrucomicrobium, Bacteriodetes, Planctomycetes, Chloroflexi, Candidate Division OP10 and Alpha-, Beta- and Gammaproteobacteria, were present in the bloom community. Diversity estimates determined from 16S rRNA gene analysis and direct cell counts and morphological identification of phytoplanktons suggested that the bloom community was dominated by members of the genera Aphanizomenon and Cylindrospermopsis, phylum Cyanobacteria. BAC-end sequencing of 37 randomly selected clones and subsequent sequence analysis provided a snapshot of the total bloom community putative metabolic activities. The sequencing of the entire inserts of seven clones (clones designated 578, 67, 142, 543, 905, 1664 and 2089) selected from BAC-end sequence studies resulted in the generation of a total of 144-kb sequence data and in the identification of 130 genes for putative proteins representing at least four phyla, Proteobacteria, Actinobacteria, Bacteroidetes and Cyanobacteria. This is the first report on a snapshot analysis of a limited metagenome of a toxic cyanobacterial freshwater bloom.
This review presents the main genera and species of the oscillatorialean cyanobacteria, that can be observed in freshwater planktic biotopes. The taxa are classified according to combined modern, ultrastructural and molecular criteria. The review is organised in form of subsequent tables.
This is the first detailed study on the occurrence of cyanobacterial toxins in India, where we studied five eutrophic, temple ponds in the vicinity of Varanasi city, Uttar Pradesh, which continuously supported blooms of Microcystis sp. for several years. Bloom material from all five ponds was sampled bi-monthly from September 2003 to August 2004. Analysis of extracts by high-performance liquid chromatography (HPLC) indicated that microcystin-RR (MC-RR) was present all year round at high concentrations (311–1540 μg/g, dry weight), posing a significant health hazard especially since all five ponds are widely used for bathing, washing, cattle drinking supply, irrigation and recreation. In addition, there was unusually low temporal variation in concentration of MC-RR in each pond, <20% variation in four out of five ponds throughout the year.Characterization of microcystin composition of several bloom samples from this study by HPLC–PDA/MS confirmed that additional microcystins were present in many of the samples. The rarely reported, MC-AR was frequently detected in bloom samples from three of the ponds (Adityanagar, Durgakund and Sankuldhara), where it typically represented 20% of the microcystin pool. MC-WR was frequently found in samples from Adityanagar and Sankuldhara, representing 5–10% of the microcystin pool. MC-LR, along with the previously unreported MC-AHar, each represented approximately 5% of the microcystin pool when present. Bloom samples from each pond had a characteristic microcystin profile, when sampled from 2003 to 2006, suggesting persistent species/strain domination.The perennial and consistent nature of the toxic Microcystis blooms in these ponds is highly unusual, in contrast to the commonly encountered temporal and spatial variation of toxigenic and non-toxigenic species. Laboratory isolates from several ponds were shown to produce microcystins, showing similar microcystin composition to the original bloom material.
Mass growths of cyanobacteria (blue–green algae), leading to the production of blooms, scums, and mats can occur in nutrient-enriched waterbodies throughout the world. Cyanobacterial mass accumulations attract the attention of water authorities and utilities, environmental and health agencies, and water-user groups, since they present water-treatment, supply, conservation, and health problems. The ability of cyanobacterial populations to produce potent toxins and annual examples of associated human and animal health problems have raised the position of cyanobacteria in the priorities for the management and protection of water quality in countries where health problems associated with the toxins have been perceived. This paper reviews the occurrence of cyanobacterial toxins in waterbodies with examples from national and local surveys. Properties of the known cyanobacterial toxins, associated health problems, and the likelihood of further toxins being discovered are discussed. The need for countermeasures to reduce or prevent the undesirable effects of cyanobacterial toxins exists at several levels. Driving forces influencing the perception of this need and the formulation of management measures include increases in eutrophication increasing demands on diminishing resources for drinking, irrigation, and leisure; and the attention of environmental and health agencies. Health hazards presented by cyanobacterial toxins and derived risk assessments are likely to increasingly influence the ranking of eutrophication problems among water quality issues in drinking and recreational waters.
1. Introduction, 35S 4. Reported incidents of cyanobacterial poisoning2. The toxins 4.1 Outbreaks associated with water contact, 37S2.1 Hepatotoxins, 35S 4.2 Poisoning associated with drinking water, 37S2.2 Neurotoxins, 36S 4.3 Potential for teratogenicity and mutagenicity from2.3 Lipopolysaccharide endotoxin, 36S drinking water, 37S3. Experimental studies of chronic administration, 5. Conclusions, 37S36S 6. References, 39S
La creciente eutrofización de los ambientes acuáticos favorece el crecimiento masivo (blooms) de algas, y poblaciones de cianobacterias, capaces de producir potentes toxinas con graves repercusiones en la salud pública y en la sanidad animal. Más del 50% de las proliferaciones masivas de cianobacterias son tóxicas. Dentro de una misma especie de cianobacteria, existen cepas que producen toxinas y otras que no las producen. Se hace necesario disponer de métodos que permitan detectar y cuantificar cianobacterias y sus toxinas. Los tradicionales bioensayos en ratón para conocer la toxicidad de una muestra sospechosa, se van sustituyendo por otros bioensayos y diversos métodos in vitro que están demostrando ser eficaces. El avance experimentado en las técnicas inmunológicas y enzimáticas ha facilitado la detección rápida de toxinas mediante el empleo de kits comerciales y no comerciales. Uno de los campos más prometedores de investigación, consiste en la determinación de las secuencias genéticas que diferencian géneros incluso cepas tóxicas, así como otras que codifican para la síntesis de toxinas; métodos que permitirán prever el desarrollo de cianobacterias o sus toxinas. El control y seguimiento de los desarrollos masivos de cianobacterias, es muy importante para garantizar la calidad del agua. El reconocimiento de los factores ambientales que influyen o desencadenan la aparición de blooms, es otro factor fundamental para la prevención y el control.
Se evaluaron las condiciones ambientales de 6 sitios a orillas del lago Titikaka (La Paz, Bolivia), encontrándose diferentes grados de eutrofización. Se emplearon 4 indicadores de evaluación rápida: macrófitas flotantes y sumergidas, fitoplancton y propiedades organolépticas, complementándolos con análisis de pH y micro-meso fauna. Se encontraron diferentes grados de afectación por el proceso eutrófico en parte debido a la influencia humana y la contaminación de los sitios de estudio. Se analizó la relevancia de cada indicador en este caso particular.
Cylindrospermopsis raciborskii is a toxic-bloom-forming cyanobacterium that is commonly found in tropical to subtropical climatic regions worldwide, but
it is also recognized as a common component of cyanobacterial communities in temperate climates. Genetic profiles of C. raciborskii were examined in 19 cultured isolates originating from geographically diverse regions of Australia and represented by two
distinct morphotypes. A 609-bp region of rpoC1, a DNA-dependent RNA polymerase gene, was amplified by PCR from these isolates with cyanobacterium-specific primers. Sequence
analysis revealed that all isolates belonged to the same species, including morphotypes with straight or coiled trichomes.
Additional rpoC1 gene sequences obtained for a range of cyanobacteria highlighted clustering of C. raciborskii with other heterocyst-producing cyanobacteria (orders Nostocales andStigonematales). In contrast, randomly amplified polymorphic DNA and short tandemly repeated repetitive sequence profiles revealed a greater
level of genetic heterogeneity amongC. raciborskii isolates than did rpoC1 gene analysis, and unique band profiles were also found among each of the cyanobacterial genera examined. A PCR test targeting
a region of therpoC1 gene unique to C. raciborskii was developed for the specific identification of C. raciborskiifrom both purified genomic DNA and environmental samples. The PCR was evaluated with a number of cyanobacterial isolates,
but a PCR-positive result was only achieved with C. raciborskii. This method provides an accurate alternative to traditional morphological identification of C. raciborskii.
This chapter focuses on the isolation and purification of Cyanobacteria. Cyanobacterial populations recognized in their natural habitat should be sampled with sterile instruments and placed in sterile containers to ensure the origin of eventual isolates. If funds permit, commercial sterile disposable scalpels, pipets, and plastic tubes are very convenient for this purpose. Only small quantities (a pea-size equivalent generally being ample) are required from habitats where macroscopic growth is visible. Sampling of endosymbiotic cyanobacteria from coralloid nodules of Cycadaceae or the stems of Gunnera can be performed as described for soil and rock-borne cyanobacteria, but other host-cyanobacteria associations might require more special treatments. To isolate cyanobacteria from lakes and ponds in which cyanobacterial growth is not visible with the eye (or even after examination with a portable microscope) it is advisable to take larger samples: 250- to 500-ml sterile screw-cap centrifuge pots, filled almost completely with sampling water, are convenient containers for transport and allow immediate concentration (by centrifugation) on arrival in the laboratory, the sampling volume generally being sufficient to isolate cyanobacteria present even in only low numbers. The origin of many cyanobacteria currently in culture is poorly characterized because little more is known about their habitat than that they were derived from a soil sample, freshwater, or marine environment, which is rather restricted information (although better than source unknown, another not uncommon description).
Some simple formulae were obtained which enable us to estimate evolutionary distances in terms of the number of nucleotide substitutions (and, also, the evolutionary rates when the divergence times are known). In comparing a pair of nucleotide sequences, we distinguish two types of differences; if homologous sites are occupied by different nucleotide bases but both are purines or both pyrimidines, the difference is called type I (or "transition" type), while, if one of the two is a purine and the other is a pyrimidine, the difference is called type II (or "transversion" type). Letting P and Q be respectively the fractions of nucleotide sites showing type I and type II differences between two sequences compared, then the evolutionary distance per site is K = -(1/2) ln [(1-2P-Q) square root of 1-2Q]. The evolutionary rate per year is then given by k = K/(2T), where T is the time since the divergence of the two sequences. If only the third codon positions are compared, the synonymous component of the evolutionary base substitutions per site is estimated by K'S = -(1/2) ln (1-2P-Q). Also, formulae for standard errors were obtained. Some examples were worked out using reported globin sequences to show that synonymous substitutions occur at much higher rates than amino acid-altering substitutions in evolution.
The 16S rRNA sequence of Trichodesmium sp. strain NIBB 1067 was determined and used for the construction of a distance tree and bootstrap analysis. The tree shows that, among the available cyanobacterial 16S rRNA sequences, Trichodesmium NIBB 1067 has Oscillatoria PCC 7515 as its closest relative, presenting 94.9% of sequence similarity with the latter strain. This is in contrast to a difference of 9 mol% G+C in mean genomic DNA base composition between the two organisms. Nevertheless, the genotypic heterogeneity presented by a number of strains assigned to the genus Oscillatoria hinders a taxonomic decision on the separate existence of the genera Trichodesmium and Oscillatoria. The sequence of the internal transcribed spacer (ITS) between the 16S and 23S rRNA genes was also determined, as a possible marker to study inter- and intraspecific variability. The ITS contains the genes coding for tRNA(Ile) and tRNA(Ala) and its total length is 547 nucleotides. In six out of eight sequenced clones, there is a duplication of 29 nucleotides, surrounding the 5' end of the tRNA(Ile).
In order to determine the nearly complete 16S rRNA gene sequences of cyanobacteria originating from nonaxenic cultures, a cyanobacterium-specific oligonucleotide probe was developed to distinguish polymerase chain reaction (PCR) products of the cyanobacterial rRNA operons from those resulting from amplification of contaminating bacteria. Using this screening method the 16S rRNA genes of four nonaxenic filamentous cyanobacterial strains belonging to the genera Leptolyngbya and Oscillatoria were cloned and sequenced. For the genus Leptolyngbya, the 16S rRNA sequence of the axenic strain PCC 73110 was also determined. Phylogenetic trees were constructed based on complete and partial sequences. The results show that the strains Leptolyngbya foveolarum Komárek 1964/112, Leptolyngbya sp. VRUC 135 Albertano 1985/1, and Leptolyngbya boryanum PCC 73110 belong to the same cluster. Strain Oscillatoria cf. corallinae SAG 8.92, which contains the rare photosynthetic pigment CU-phycoerythrin, is not closely related to other CU-phycoerythrin-containing cyanobacteria. Oscillatoria agardhii CYA 18, which is a representative of planktonic Oscillatoria species that form toxic blooms in Norwegian inland waters, has no close relatives in the tree.
Cyanobacteria, broadly classified as oxygenic phototrophs containing chlorophyll- a and accessory pigments, are among the oldest life forms on earth. They may be unicellular, colonial or filamentous, with cell sizes varying from less than 2 μm to 40 μm in diameter. They may live as symbionts with plants and fungi, in the benthos or in the water column. Cyanobacteria have a cosmopolitan distribution and are common in all kinds of habitats, including Antarctic lakes, thermal springs, arid deserts and tropical acidic soils. Most commonly they are known for their existence as planktonic members of the water column in marine and freshwater environments.
Cyanobacteria have special adaptations such as; nitrogen fixation, ability to regulate buoyancy, light harvesting pigments, and differentiated cell types for reproduction or resting, allowing them an advantage over many competitors. Anthropogenic factors such as increased nutrient loading in freshwater and coastal environments, have lead to the occurrence of cyanobacterial blooms world wide. While blooms are unpleasant both visually and due to released odour and taste factors, certain metabolites produced by some cyanobacterial species pose a more serious problem [1]. Due to their adverse effects on higher organisms, these compounds have been labelled as ‘toxins’ and from here on are referred to as such. The chemical structures of many cyanotoxins and their adverse effect on animals have been elucidated and recently reviewed [2]. However, the actual physiological function and ecological regulation of cyanobacterial toxins remain largely a mystery.
Studies on regulation and function of cyanotoxins are interrelated and have focused on their effect on the local ecosystem, as well as the effect of environmental parameters on toxin production. Hypotheses regarding the ecophysiology of the cyanotoxins, stemming from such studies, are discussed in this review. With advances in molecular biology, it has more recently been possible …