The increased proliferation of benthic marine cyanobacteria of the Lyngbya genus in many tropical and subtropical systems worldwide is a concern due to the detrimental impacts these blooms can have on ecosystems, local economies and public health. While increasing nutrient loads from anthropogenic sources/activities has been hypothesised as the main cause, evidence to support this is limited. This hypothesis was explored by investigating blooms of the toxic, benthic cyanobacterium Lyngbya majuscula in a sub-tropical shallow coastal embayment (Moreton Bay) in southeast Queensland, Australia—where blooms have increased in frequency and severity. More specifically, the thesis aimed to investigate the role of nutrients in the physiology and growth dynamics of L. majuscula in Moreton Bay through examination of three main research questions. Examination of the spatial and temporal variations in the growth and nutritional status of L. majuscula in Moreton Bay (Research Question 1) was investigated by tracking natural summer blooms in northeastern Moreton Bay (Deception Bay) over two successive years. Detailed field observations, extensive biomass and tissue nutrient sampling (every 10–14 days) and a three-dimensional model were used to map the change in areal extent, biomass and tissue nutrients over the course of the blooms. The results demonstrated the innate ability of L. majuscula to rapidly spread and generate massive amounts of biomass, with the peak biomass calculated at 5057 tww in the 2005–2006 and 10 213 tww in the 2006–2007 seasons. A sequence of phases showing differing appearance, biomass growth and tissue nutrient changes were identified and documented. The role of nutrients (individually and collectively) in the enhancement of L. majuscula growth (Research Question 2) was investigated using a combination of comprehensive laboratory experiments (filament growth, 14C-bicarbonate uptake rate and biomass increase) and in-situ field experiments. Addition of nutrients to the water column were shown to promote prolific L. majuscula growth in the laboratory; this was confirmed in field experiments at two locations in Moreton Bay—showing nutrients can be a major causal factor in bloom formation. Additions of phosphorus (macronutrient) and iron (required for photosynthesis and nitrogen-fixation) caused the greatest stimulation of L. majuscula in both laboratory and field experiments. The form of iron was shown to be important —organically complexed iron (FeEDTA) was substantially more effective in promoting L. majuscula growth under laboratory conditions than inorganic iron (FeCl3). This is important as FeEDTA mirrors the naturally occurring iron organic complexes (which increase the solubility of iron) in waters from the region. The effect of nitrogen additions was more complex—likely due to the capacity of L. majuscula to fix atmospheric nitrogen reducing reliance on an inorganic nitrogen source. In the high light conditions experienced in this study, L. majuscula appeared to acquire nitrogen: (i) directly from the dissolved inorganic nitrogen in the water column—evidenced by a positive response to the nitrogen treatments; and, (ii) through enhanced nitrogen-fixation rates when iron and/or phosphorus were added in the absence of nitrogen—inferred from a substantial increase in the total nitrogen content of the L. majuscula biomass (nitrogen-fixation was not measured directly). The main sources of naturally occurring nutrients likely to promote L. majuscula blooms in Moreton Bay (Research Question 3) were investigated using laboratory experiments, soil and water analyses, and GIS-based modelling. The potential for groundwater/surfacewater from different vegetation, soils, geology and landuses within the study area catchments to stimulate L. majuscula response (14C-bicarbonate uptake rate) was tested in laboratory bioassays. Areas with acid sulfate soils (ASS), Melaleuca vegetation, pine plantations and Casuarina on ASS all had waters that enhanced L. majuscula growth. To investigate causal agents, bioassay response data and water analyses were subject to multiple regression and correlation analysis; this confirmed the importance of iron, phosphorus and nitrogen to L. majuscula growth and the roles of low pH and dissolved organic carbon, the latter two appearing to influence the chemical state and enhance the solubility of nutrients to L. majuscula. This information was incorporated into a GIS-based model to identify areas of hazard which were most likely to supply/export nutrients to Moreton Bay. These hazard maps, with further local verification, will be used as planning and decision support tools to assist government and landuse managers to limit the mobilisation and transport of key nutrients to potential bloom sites. The results from this thesis demonstrate that a precautionary approach to limit phosphorus, iron, nitrogen and dissolved organic carbon to waterways is necessary; otherwise the magnitude of L. majuscula blooms is likely to increase in Moreton Bay as coastal development intensifies with the predicted population increase. The thesis findings provide strong support for the hypothesised link between nutrients and the increased proliferation of Lyngbya and other benthic cyanobacteria blooms and are likely to be applicable to other areas where environmental conditions are suitable for their growth.

The increased proliferation of benthic marine cyanobacteria of the Lyngbya genus in many tropical and subtropical systems worldwide is a concern due to the detrimental impacts these blooms can have on ecosystems, local economies and public health. While increasing nutrient loads from anthropogenic sources/activities has been hypothesised as the main cause, evidence to support this is limited. This hypothesis was explored by investigating blooms of the toxic, benthic cyanobacterium Lyngbya majuscula in a sub-tropical shallow coastal embayment (Moreton Bay) in southeast Queensland, Australia—where blooms have increased in frequency and severity. More specifically, the thesis aimed to investigate the role of nutrients in the physiology and growth dynamics of L. majuscula in Moreton Bay through examination of three main research questions. Examination of the spatial and temporal variations in the growth and nutritional status of L. majuscula in Moreton Bay (Research Question 1) was investigated by tracking natural summer blooms in northeastern Moreton Bay (Deception Bay) over two successive years. Detailed field observations, extensive biomass and tissue nutrient sampling (every 10–14 days) and a three-dimensional model were used to map the change in areal extent, biomass and tissue nutrients over the course of the blooms. The results demonstrated the innate ability of L. majuscula to rapidly spread and generate massive amounts of biomass, with the peak biomass calculated at 5057 tww in the 2005–2006 and 10 213 tww in the 2006–2007 seasons. A sequence of phases showing differing appearance, biomass growth and tissue nutrient changes were identified and documented. The role of nutrients (individually and collectively) in the enhancement of L. majuscula growth (Research Question 2) was investigated using a combination of comprehensive laboratory experiments (filament growth, 14C-bicarbonate uptake rate and biomass increase) and in-situ field experiments. Addition of nutrients to the water column were shown to promote prolific L. majuscula growth in the laboratory; this was confirmed in field experiments at two locations in Moreton Bay—showing nutrients can be a major causal factor in bloom formation. Additions of phosphorus (macronutrient) and iron (required for photosynthesis and nitrogen-fixation) caused the greatest stimulation of L. majuscula in both laboratory and field experiments. The form of iron was shown to be important —organically complexed iron (FeEDTA) was substantially more effective in promoting L. majuscula growth under laboratory conditions than inorganic iron (FeCl3). This is important as FeEDTA mirrors the naturally occurring iron organic complexes (which increase the solubility of iron) in waters from the region. The effect of nitrogen additions was more complex—likely due to the capacity of L. majuscula to fix atmospheric nitrogen reducing reliance on an inorganic nitrogen source. In the high light conditions experienced in this study, L. majuscula appeared to acquire nitrogen: (i) directly from the dissolved inorganic nitrogen in the water column—evidenced by a positive response to the nitrogen treatments; and, (ii) through enhanced nitrogen-fixation rates when iron and/or phosphorus were added in the absence of nitrogen—inferred from a substantial increase in the total nitrogen content of the L. majuscula biomass (nitrogen-fixation was not measured directly). The main sources of naturally occurring nutrients likely to promote L. majuscula blooms in Moreton Bay (Research Question 3) were investigated using laboratory experiments, soil and water analyses, and GIS-based modelling. The potential for groundwater/surfacewater from different vegetation, soils, geology and landuses within the study area catchments to stimulate L. majuscula response (14C-bicarbonate uptake rate) was tested in laboratory bioassays. Areas with acid sulfate soils (ASS), Melaleuca vegetation, pine plantations and Casuarina on ASS all had waters that enhanced L. majuscula growth. To investigate causal agents, bioassay response data and water analyses were subject to multiple regression and correlation analysis; this confirmed the importance of iron, phosphorus and nitrogen to L. majuscula growth and the roles of low pH and dissolved organic carbon, the latter two appearing to influence the chemical state and enhance the solubility of nutrients to L. majuscula. This information was incorporated into a GIS-based model to identify areas of hazard which were most likely to supply/export nutrients to Moreton Bay. These hazard maps, with further local verification, will be used as planning and decision support tools to assist government and landuse managers to limit the mobilisation and transport of key nutrients to potential bloom sites. The results from this thesis demonstrate that a precautionary approach to limit phosphorus, iron, nitrogen and dissolved organic carbon to waterways is necessary; otherwise the magnitude of L. majuscula blooms is likely to increase in Moreton Bay as coastal development intensifies with the predicted population increase. The thesis findings provide strong support for the hypothesised link between nutrients and the increased proliferation of Lyngbya and other benthic cyanobacteria blooms and are likely to be applicable to other areas where environmental conditions are suitable for their growth.

Thesis (Ph. D.)--Oregon State University, 2009. / Printout. Includes bibliographical references (p. 153-160) Also available on the World Wide Web.

abstract: Some cyanobacteria can generate hydrogen (H2) under certain physiological conditions and are considered potential agents for biohydrogen production. However, they also present low amounts of H2 production, a reaction reversal towards H2 consumption, and O2 sensitivity. Most attempts to improve H2 production have involved genetic or metabolic engineering approaches. I used a bio-prospecting approach instead to find novel strains that are naturally more apt for biohydrogen production. A set of 36, phylogenetically diverse strains isolated from terrestrial, freshwater and marine environments were probed for their potential to produce H2 from excess reductant. Two distinct patterns in H2 production were detected. Strains displaying Pattern 1, as previously known from Synechocystis sp. PCC 6803, produced H2 only temporarily, reverting to H2 consumption within a short time and after reaching only moderately high H2 concentrations. By contrast, Pattern 2 cyanobacteria, in the genera Lyngbya and Microcoleus, displayed high production rates, did not reverse the direction of the reaction and reached much higher steady-state H2 concentrations. L. aestuarii BL J, an isolate from marine intertidal mats, had the fastest production rates and reached the highest steady-state concentrations, 15-fold higher than that observed in Synechocystis sp. PCC 6803. Because all Pattern 2 strains originated in intertidal microbial mats that become anoxic in dark, it was hypothesized that their strong hydrogenogenic capacity may have evolved to aid in fermentation of the photosynthate. When forced to ferment, these cyanobacteria display similarly desirable characteristics of physiological H2 production. Again, L. aestuarii BL J had the fastest specific rates and attained the highest H2 concentrations during fermentation, which proceeded via a mixed-acid pathway to yield acetate, ethanol, lactate, H2, CO2 and pyruvate. The genome of L. aestuarii BL J was sequenced and bioinformatically compared to other cyanobacterial genomes to ascertain any potential genetic or structural basis for powerful H2 production. The association hcp exclusively in Pattern 2 strains suggests its possible role in increased H2 production. This study demonstrates the value of bioprospecting approaches to biotechnology, pointing to the strain L. aestuarii BL J as a source of useful genetic information or as a potential platform for biohydrogen production. / Dissertation/Thesis / Ph.D. Molecular and Cellular Biology 2013

Pahayokolides A-D are cytotoxic cyclic polypeptides produced by the freshwater cyanobacterium Lyngbya sp. strain 15-2 that possess an unusual β-amino acid, 3-amino-2,5,7,8-tetrahydroxy-10-methylundecanoic acid (Athmu). The absolute configuration of pahayokolides A-D was determined using advanced Marfey’s method. It was also confirmed that a pendant N-acetyl-N-methyl leucine moiety in pahayokolide A was absent in pahayokolides B and pahayokolides C-D were conformers of pahayokolide A. Feeding experiments indicated that the biosynthesis of the Athmu sidechain arises from leucine or α-ketoisovalerate, however could not be further extended by three rounds of condensation with malonate units. Putative four peptide and one unique polyketide synthetases in Lyngbya sp. strain 15-2 were identified by using a PCR method and degenerate primers derived from conserved core sequences of known NRPSs and PKSs. Identification of one unique KS domain conflicted with the logic rule that the long side chain of Athmu was assembled by three rounds of ketide extensions if PKSs were involved. A gene cluster (pah) encoding a peptide synthetase putatively producing pahayokolide was cloned, partially sequenced and characterized. Seven modules of the non-ribosomal peptide synthetase (NRPS) were identified. Ten additional opening reading frames (ORFs) were found, responsible for peptide resistance, transport and degradation. Although the predicted substrate specificities of NRPS agreed with the structure of pahayokolide A partially, the disagreement could be explained. However, no PKS gene was found in the pah gene cluster.

No description available.

No description available.

No description available.

Les proliférations nuisibles de la cyanobactérie filamenteuse benthique Lyngbya wollei qui forme des tapis déposés sur les sédiments ont augmenté en fréquence au cours des 30 dernières années dans les rivières, lacs et sources de l'Amérique du Nord. Lyngbya wollei produit des neurotoxines et des composés organiques volatils (géosmin, 2-méthylisobornéol) qui ont des répercussions sur la santé publique de même que des impacts d'ordre socioéconomiques. Cette cyanobactérie est considérée comme un habitat et une source de nourriture de piètre qualité pour les invertébrés en raison de sa gaine robuste et de sa production de toxines. Les proliférations de L. wollei ont été observées pour la première fois en 2005 dans le fleuve Saint-Laurent (SLR; Québec, Canada). Nous avons jugé important de déterminer sa distribution sur un tronçon de 250 km afin d'élaborer des modèles prédictifs de sa présence et biomasse en se basant sur les caractéristiques chimiques et physiques de l'eau. Lyngbya wollei était généralement observé en aval de la confluence de petits tributaires qui irriguent des terres agricoles. L’écoulement d’eaux enrichies à travers la végétation submergée se traduisait par une diminution de la concentration d’azote inorganique dissous (DIN), alors que les concentrations de carbone organique dissous (DOC) et de phosphore total dissous (TDP) demeuraient élevées, produisant un faible rapport DIN :TDP. Selon nos modèles, DOC (effet positif), TP (effet négatif) et DIN :TDP (effet négatif) sont les variables les plus importantes pour expliquer la répartition de cette cyanobactérie. La probabilité que L. wollei soit présent dans le SLR a été prédite avec exactitude dans 72 % à 92 % des cas pour un ensemble de données indépendantes. Nous avons ensuite examiné si les conditions hydrodynamiques, c'est-à-dire le courant généré par les vagues et l'écoulement du fleuve, contrôlent les variations spatiales et temporelles de biomasse de L. wollei dans un grand système fluvial. Nous avons mesuré la biomasse de L. wollei ainsi que les variables chimiques, physiques et météorologiques durant trois ans à 10 sites le long d'un gradient d'exposition au courant et au vent dans un grand (148 km2) lac fluvial du SLR. L'exposition aux vagues et la vitesse du courant contrôlaient les variations de biomasses spatiales et temporelles. La biomasse augmentait de mai à novembre et persistait durant l'hiver. Les variations interannuelles étaient contrôlées par l'écoulement de la rivière (niveau d'eau) avec la crue printanière qui délogeait les tapis de l'année précédente. Les baisses du niveau d'eau et l'augmentation de l'intensité des tempêtes anticipées par les scénarios de changements climatiques pourraient accroître la superficie colonisée par L. wollei de même que son accumulation sur les berges. Par la suite, nous avons évalué l'importance relative de L. wollei par rapport aux macrophytes et aux épiphytes. Nous avons examiné l'influence structurante de l'échelle spatiale sur les variables environnementales et la biomasse de ces producteurs primaires (PP) benthiques. Nous avons testé si leur biomasse reflétait la nature des agrégats d'habitat basées sur l'écogéomorphologie ou plutôt le continuum fluvial. Pour répondre à ces deux questions, nous avons utilisé un design à 3 échelles spatiales dans le SLR: 1) le long d'un tronçon de 250 km, 2) entre les lacs fluviaux localisés dans ce tronçon, 3) à l'intérieur de chaque lac fluvial. Les facteurs environnementaux (conductivité et TP) et la structure spatiale expliquent 59% de la variation de biomasse des trois PP benthiques. Spécifiquement, les variations de biomasses étaient le mieux expliquées par la conductivité (+) pour les macrophytes, par le ratio DIN:TDP (+) et le coefficient d'extinction lumineuse (+) pour les épiphytes et par le DOC (+) et le NH4+ (-) pour L. wollei. La structure spatiale à l'intérieur des lacs fluviaux était la plus importante composante spatiale pour tous les PP benthiques, suggérant que les effets locaux tels que l'enrichissement par les tributaire plutôt que les gradients amont-aval déterminent la biomasse de PP benthiques. Donc, la dynamique des agrégats d'habitat représente un cadre général adéquat pour expliquer les variations spatiales et la grande variété de conditions environnementales supportant des organismes aquatiques dans les grands fleuves. Enfin, nous avons étudié le rôle écologique des tapis de L. wollei dans les écosystèmes aquatiques, en particulier comme source de nourriture et refuge pour l'amphipode Gammarus fasciatus. Nous avons offert aux amphipodes un choix entre des tapis de L. wollei et soit des chlorophytes filamenteuses ou un tapis artificiel de laine acrylique lors d'expériences en laboratoire. Nous avons aussi reconstitué la diète in situ des amphipodes à l'aide du mixing model (d13C et δ15N). Gammarus fasciatus choisissait le substrat offrant le meilleur refuge face à la lumière (Acrylique>Lyngbya=Rhizoclonium>Spirogyra). La présence de saxitoxines, la composition élémentaire des tissus et l'abondance des épiphytes n'ont eu aucun effet sur le choix de substrat. Lyngbya wollei et ses épiphytes constituaient 36 et 24 % de l'alimentation in situ de G. fasciatus alors que les chlorophytes, les macrophytes et les épiphytes associées représentaient une fraction moins importante de son alimentation. Les tapis de cyanobactéries benthiques devraient être considérés comme un bon refuge et une source de nourriture pour les petits invertébrés omnivores tels que les amphipodes. / Harmful proliferations of the filamentous cyanobacterium L. wollei forming conspicuous benthic mats on the bottom sediment have been reported with increasing frequency in the last 30 years in rivers, lakes, and springs in North America. It is a known producer of neurotoxins and volatile organic compounds (geosmin, 2-methylisoborneol) thus exerting socioeconomic and public health impacts. Lyngbya wollei is also considered a poor nutritional source for invertebrates because of its robust sheath and toxin production. Proliferation of L. wollei in St. Lawrence River (SLR; Quebec, Canada) was first noticed in 2005. We deemed important to determine its distribution over a 250 km stretch of the SLR to elaborate predictive models of its presence and biomass based on chemical and physical characteristics. Lyngbya wollei was generally found downstream of the inflow tributaries draining farmlands. As enriched waters flowed slowly through submerged vegetation, dissolved inorganic nitrogen (DIN) concentration dropped but dissolved organic carbon (DOC) and total dissolved phosphorus (TDP) remained high, leading to a low DIN:TDP ratio. Models identified DOC (positive effect), TP (negative effect), and DIN:TDP (negative effect) as the most important variables explaining L. wollei distribution. The risk of L. wollei occurrence in the SLR was correctly forecasted in 72%-92% of all cases with an independent data set. We then examined if hydrodynamic conditions, namely currents generated by waves and river flow, control spatial and temporal variations of L. wollei biomass in a large river system. We measured L. wollei biomass together with meteorological, physical, and chemical variables over three years at 10 sites along a gradient of exposure to current and wind in a large (148 km2) fluvial lake of SLR. Wave exposure and current velocity controlled spatial and temporal biomass variations. Biomass increased from May to November and persisted during winter. Interannual variations were primarily controlled by river flow (water level) with spring discharge dislodging mats from the previous year. As anticipated under climate change scenarios, drops in water level and rising storm intensity may lead to an increase in the areas colonized by L. wollei, together with more frequent episodes of mat disruption and beach fouling. Additionally, we evaluated the relative importance of L. wollei with respect to macrophytes and epiphytes. We assessed the influence of the spatial scale in structuring environmental variables and biomass of these benthic primary producers (PP). We also test to which extent their biomass reflected the nature of patches based on ecogeomorphology or the river continuum. To address these two questions, we used a nested design at 3 spatial scales within the SLR: 1) along a 250-km-long upstream-downstream river stretch, 2) among three fluvial lakes located within that river stretch and 3) within each fluvial lake. Environmental factors (conductivity and TP) and spatial structure together explained 59% of the variability in biomass of all three benthic PP. Spatial variability of biomass was best explained by conductivity (+) for macrophytes, DIN:TDP ratio (+) and water extinction coefficient (+) for epiphytes and DOC (+) and NH4+ (-) for L. wollei mats. Within-lake structure was the most important spatial component for all benthic PP, suggesting that local effects, such as enrichment by the inflow of tributaries, rather than upstream-downstream gradients, determined the biomass and composition of benthic PP. Therefore patch dynamics represents a general framework which adequately covers the spatial variability and wide variety of environmental conditions experienced by aquatic organisms found in large rivers. Finally, we investigated the ecological role of L. wollei mats in aquatic ecosystems, especially as a food source and shelter for the amphipod Gammarus fasciatus. We offered amphipods a choice between mats of L. wollei and either chlorophytes or an artificial mat made of acrylic wool in laboratory experiment. Moreover, we reconstructed in situ amphipod diet using mixing model (δ13C and δ15N). Gammarus fasciatus selected the substratum offering the best light refuge (Acrylic > Lyngbya = Rhizoclonium > Spirogyra). Presence of saxitoxins, tissue elemental composition and epiphyte abundance had no significant effect on substratum choice. Lyngbya wollei and its epiphytes constituted 36 and 24% of the in situ diet of G. fasciatus whereas chlorophytes, macrophytes and associated epiphytes represented a less important fraction of its diet. Benthic cyanobacterial mats should be considered a good shelter and food source for small omnivorous invertebrates such as amphipods.