Diazotrophic Filamentous Cyanobacteria Research Articles

Cobalt as a potential limiting factor for heterocyst frequency in nitrogen‐limited Aphanizomenon and Dolichospermum: Evidence from experimental and field studies

Abstract Evidence suggests that low cobalt (Co) may limit heterocyst frequency (HF) in filamentous diazotrophic cyanobacteria. Therefore, the effect of three Co concentrations on HF in four species in the genera Aphanizomenon and Dolichospermum was examined in N‐depleted (N was not added) culture. Long‐term HF data were also analysed in experimentally fertilised N‐limited Lake 227. HF ranges and means at 0.17, 17, and 170 nmol Co/L were 4.1%–5.7% (4.6%), 5.4%–7.4% (6.4%), and 5.9%–9.3% (7.4%), respectively, after 11 days of growth in batch cultures suggesting that Co plays a role in regulating HF. HF at 0.17 nmol/L was significantly different from HF at the two higher concentrations in all species implicating Co in heterocyst differentiation. However, growth rates and final biovolume yields were not significantly affected by Co in any of the species. Applying the ratio of total phosphorus (TP)/Co in culture media (106) at 0.17 nmol Co/L to an N‐limited system implies that dissolved Co would have to be less than 1 pmol/L to severely limit HF at 1 μmol TP/L. This Co concentration is much lower than dissolved Co in Lake 227 during an Aphanizomenon skujae bloom with 1 μmol TP/L in 2017. Furthermore, Co increased from 0.7 to 2.0 nmol/L simultaneously increasing with HF and cyanobacteria biomass suggesting that Co probably did not limit HF and bloom biomass in Lake 227. Mean summer HFs in Lake 227 during 2000–2020 were 3.4% (epilimnion) and 4.0% (metalimnion), similar to HF at the lowest Co in the cultures. However, HF was significantly higher after 2015 following a shift in dominant bloom species from A. schindlerii to the smaller A. skujae. Calculations suggest that the decrease in cell size may have prompted a higher HF in A. skujae in order to fix an amount of N per unit biomass similar to that fixed by the larger A. schindlerii. Thus, two previously unidentified factors, Co and cell size, can be added to the list of factors that affect HF.

Whole-genome sequence of the filamentous diazotrophic cyanobacterium Tolypothrix sp. PCC 7712 and its comparison with non-diazotrophic Tolypothrix sp. PCC 7601.

Cyanobacteria are highly promising microorganisms in forthcoming biotechnologies. Besides the systematic development of molecular tools for genetic engineering, the design of chassis strains and novel reactor concepts are in focus. The latter includes capillary biofilm reactors (CBR), which offer a high surface area-to-volume ratio and very high cell densities. In this context, Tolypothrix sp. PCC 7712 was found to be highly suited for this reactor system due to maximal surface coverage, extraordinarily strong biofilm attachment, and high biomass formation. Here, we provide the genome sequence of Tolypothrix sp. PCC 7712 to potentially allow targeted strain engineering. Surprisingly, it was almost identical to an available incomplete genome draft of Tolypothrix sp. PCC 7601. Thus, we completely sequenced this strain as well and compared it in detail to strain PCC 7712. Comparative genome analysis revealed 257 and 80 unique protein-coding sequences for strains PCC 7601 and PCC 7712, respectively. Clustering genomes based on average nucleotide identity (ANI) and 16S rRNA homology showed 99.98% similarity and only minor distance, respectively, between the two strains in contrast to 21 other cyanobacterial genomes. Despite these high similarities, both strains differ in the ability to fix atmospheric nitrogen and show specific sequence variations, which are discussed in the paper.

A tRNA t6A modification system contributes to the sensitivity towards the toxin β-N-methylamino-L-alanine (BMAA) in the cyanobacterium Anabaena sp. PCC 7120.

Cyanobacteria are oxygen-evolving photosynthetic autotrophs essential for nutrient cycling in the environment. They possess multiple control mechanisms for their cellular activities in order to adapt to the environment. While protein translation is essential for cell survival and adaptation, the regulation and the flexibility of this process are poorly understood in cyanobacteria. β-N-methylamino-L-alanine (BMAA), an amino acid analog proposed as an environmental neurotoxin, is highly toxic to the filamentous diazotrophic cyanobacterium Anabaena PCC 7120. In this study, through genetic analysis of BMAA-resistant mutants, we demonstrate that the system responsible for modification of ANN-decoding tRNAs with N(6)-threonylcarbamoyl adenosine (t6A) is involved in BMAA sensitivity through the control of translation. Both BMAA and inactivation of the t6A biosynthesis pathway affect translational fidelity and ribosome assembly. However, the two factors display either additive effects on translational elongation, or attenuate each other over translational fidelity or the resistance/sensitivity to antibiotics that inhibit different steps of the translational process. BMAA has a broad effect on translation and transcription, and once BMAA enters the cells, the presence of the t6A biosynthesis pathway increases the sensitivity of the cells towards this toxin. BMAA-resistant mutants screening is an effective method for getting insight into the toxic mechanisms of BMAA. In addition, BMAA is a useful tool for probing translational flexibility of cyanobacteria, and the characterization of the corresponding resistant mutants should help us to reveal translational mechanism allowing cyanobacteria to adapt to changing environments.

Transcriptional responses of Trichodesmium to natural inverse gradients of Fe and P availability

The filamentous diazotrophic cyanobacterium Trichodesmium is responsible for a significant fraction of marine di-nitrogen (N2) fixation. Growth and distribution of Trichodesmium and other diazotrophs in the vast oligotrophic subtropical gyres is influenced by iron (Fe) and phosphorus (P) availability, while reciprocally influencing the biogeochemistry of these nutrients. Here we use observations across natural inverse gradients in Fe and P in the North Atlantic subtropical gyre (NASG) to demonstrate how Trichodesmium acclimates in situ to resource availability. Transcriptomic analysis identified progressive upregulation of known iron-stress biomarker genes with decreasing Fe availability, and progressive upregulation of genes involved in the acquisition of diverse P sources with decreasing P availability, while genes involved in N2 fixation were upregulated at the intersection under moderate Fe and P availability. Enhanced N2 fixation within the Fe and P co-stressed transition region was also associated with a distinct, consistent metabolic profile, including the expression of alternative photosynthetic pathways that potentially facilitate ATP generation required for N2 fixation with reduced net oxygen production. The observed response of Trichodesmium to availability of both Fe and P supports suggestions that these biogeochemically significant organisms employ unique molecular, and thus physiological responses as adaptations to specifically exploit the Fe and P co-limited niche they construct.

Nitrogen fixation estimates for the Baltic Sea indicate high rates for the previously overlooked Bothnian Sea

Dense blooms of diazotrophic filamentous cyanobacteria are formed every summer in the Baltic Sea. We estimated their contribution to nitrogen fixation by combining two decades of cyanobacterial biovolume monitoring data with recently measured genera-specific nitrogen fixation rates. In the Bothnian Sea, estimated nitrogen fixation rates were 80 kt N year−1, which has doubled during recent decades and now exceeds external loading from rivers and atmospheric deposition of 69 kt year−1. The estimated contribution to the Baltic Proper was 399 kt N year−1, which agrees well with previous estimates using other approaches and is greater than the external input of 374 kt N year−1. Our approach can potentially be applied to continuously estimate nitrogen loads via nitrogen fixation. Those estimates are crucial for ecosystem adaptive management since internal nitrogen loading may counteract the positive effects of decreased external nutrient loading.

Reconstruction of a regulated two-cell metabolic model to study biohydrogen production in a diazotrophic cyanobacterium Anabaena variabilis ATCC 29413.

Anabaena variabilis is a diazotrophic filamentous cyanobacterium that differentiates to heterocysts and produces hydrogen as a byproduct. Study on metabolic interactions of the two differentiated cells provides a better understanding of its metabolism especially for improving hydrogen production. To this end, a genome-scale metabolic model for Anabaena variabilis ATCC 29413, iAM957, was reconstructed and evaluated in this research. Then, the model and transcriptomic data of the vegetative and heterocyst cells were applied to construct a regulated two-cell metabolic model. The regulated model improved prediction for biomass in high radiation levels. The regulated model predicts that heterocysts provide an oxygen-free environment and then, this model was used to find strategies for improving hydrogen production in heterocysts. The predictions indicate that the removal of uptake hydrogenase improves hydrogen production which is consistent with previous empirical research. Furthermore, the regulated model proposed activation of some reactions to provide redox cofactors which are required for improving hydrogen production up to 60% by bidirectional hydrogenase.

Multiscale Multiobjective Systems Analysis (MiMoSA): an advanced metabolic modeling framework for complex systems

In natural environments, cells live in complex communities and experience a high degree of heterogeneity internally and in the environment. Even in ‘ideal’ laboratory environments, cells can experience a high degree of heterogeneity in their environments. Unfortunately, most of the metabolic modeling approaches that are currently used assume ideal conditions and that each cell is identical, limiting their application to pure cultures in well-mixed vessels. Here we describe our development of Multiscale Multiobjective Systems Analysis (MiMoSA), a metabolic modeling approach that can track individual cells in both space and time, track the diffusion of nutrients and light and the interaction of cells with each other and the environment. As a proof-of concept study, we used MiMoSA to model the growth of Trichodesmium erythraeum, a filamentous diazotrophic cyanobacterium which has cells with two distinct metabolic modes. The use of MiMoSA significantly improves our ability to predictively model metabolic changes and phenotype in more complex cell cultures.

Limited response of a spring bloom community inoculated with filamentous cyanobacteria to elevated temperature and pCO2

Abstract Temperature and CO2 levels are projected to increase in the future, with consequences for carbon and nutrient cycling in brackish environments, such as the Baltic Sea. Moreover, filamentous cyanobacteria are predicted to be favored over other phytoplankton groups under these conditions. Under a 12-day outdoor experiment, we examined the effect on a natural phytoplankton spring bloom community of elevated temperature (from 1°C to 4°C) and elevated pCO2 (from 390 to 970 μatm). No effects of elevated pCO2 or temperature were observed on phytoplankton biovolumes, but a significantly higher photosystem II activity was observed at elevated temperature after 9 days. In addition, three species of diazotrophic filamentous cyanobacteria were inoculated to test their competitive capacity under spring bloom conditions. The toxic cyanobacterium Nodularia spumigena exhibited an average specific growth rate of 0.10 d−1 by the end of the experiment, indicating potential prevalence even during wintertime in the Baltic Sea. Generally, none of the inoculated cyanobacteria species were able to outcompete the natural phytoplankton species at temperatures ≤4°C. No direct effects were found on heterotrophic bacteria. This study demonstrates the highly efficient resistance towards short-term (12 days) changes in abiotic factors by the natural Baltic Sea spring bloom community.

Spectral microscopic imaging of heterocysts and vegetative cells in two filamentous cyanobacteria based on spontaneous Raman scattering and photoluminescence by 976 nm excitation

Photosynthetic pigment-protein complexes are highly concentrated in thylakoid membranes of chloroplasts and cyanobacteria that emit strong autofluorescence (mainly 600–800 nm). In Raman scattering microscopy that enables imaging of pigment concentrations of thylakoid membranes, near infrared laser excitation at 1064 nm or visible laser excitation at 488–532 nm has been often employed in order to avoid the autofluorescence. Here we explored a new approach to Raman imaging of thylakoid membranes by using excitation wavelength of 976 nm. Two types of differentiated cells, heterocysts and vegetative cells, in two diazotrophic filamentous cyanobacteria, Anabaena variabilis, and Rivularia M-261, were characterized. Relative Raman scattering intensities of phycobilisomes of the heterocyst in comparison with the nearest vegetative cells of Rivularia remained at a significantly higher level than those of A. variabilis. It was also found that the 976 nm excitation induces photoluminescence around 1017–1175 nm from the two cyanobacteria, green alga (Parachlorella kessleri) and plant (Arabidopsis thaliana). We propose that this photoluminescence can be used as an index of concentration of chlorophyll a that has relatively small Raman scattering cross-sections. The Rivularia heterocysts that we analyzed were clearly classified into at least two subgroups based on the Chla-associated photoluminescence and carotenoid Raman bands, indicating two physiologically distinct states in the development or aging of the terminal heterocyst.

Trichodesmium and other planktonic cyanobacteria in New Caledonian waters (SW tropical Pacific) during an El Niño episode

Contributions of filamentous and picoplanktonic cyanobacteria to the 26 phytoplankton community structure were examined in New Caledonian waters during the 2001-27 2003 El Nino period at three stations. Morphometric characteristics of diazotrophic filamentous 28 cyanobacteria are given as well as the seasonal and inter-annual variations of their surface areas 29 and integrated abundances. Trichodesmium tenue and T. thiebautii were the dominant species 30 followed by T. erythraeum, altogether accounting for more than 51-80 % of the biomass of the 31 free-living filamentous cyanobacteria. Katagnymene spp. accounted for a smaller percentage 32 (<13.8 % at ocean stations, <3.6 % in the lagoon). R. intracellularis biomass was relatively small 33 (<1 % of total surface area and volume of Trichodesmium trichomes) with the highest 34

Ocean acidification and desalination: climate-driven change in a Baltic Sea summer microplanktonic community

Helcom scenario modelling suggests that the Baltic Sea, one of the largest brackish-water bodies in the world, could expect increased precipitation (decreased salinity) and increased concentration of atmospheric CO2 over the next 100 years. These changes are expected to affect the microplanktonic food web, and thereby nutrient and carbon cycling, in a complex and possibly synergistic manner. In the Baltic Proper, the extensive summer blooms dominated by the filamentous cyanobacteria Aphanizomenon sp., Dolichospermum spp. and the toxic Nodularia spumigena contribute up to 30% of the yearly new nitrogen and carbon exported to the sediment. In a 12 days outdoor microcosm experiment, we tested the combined effects of decreased salinity (from 6 to 3) and elevated CO2 concentrations (380 and 960 µatm) on a natural summer microplanktonic community, focusing on diazotrophic filamentous cyanobacteria. Elevated pCO2 had no significant effects on the natural microplanktonic community except for higher biovolume of Dolichospermum spp. and lower biomass of heterotrophic bacteria. At the end of the experimental period, heterotrophic bacterial abundance was correlated to the biovolume of N. spumigena. Lower salinity significantly affected cyanobacteria together with biovolumes of dinoflagellates, diatoms, ciliates and heterotrophic bacteria, with higher biovolume of Dolichospermum spp. and lower biovolume of N. spumigena, dinoflagellates, diatoms, ciliates and heterotrophic bacteria in reduced salinity. Although the salinity effects on diatoms were apparent, they could not clearly be separated from the influence of inorganic nutrients. We found a clear diurnal cycle in photosynthetic activity and pH, but without significant treatment effects. The same diurnal pattern was also observed in situ (pCO2, pH). Thus, considering the Baltic Proper, we do not expect any dramatic effects of increased pCO2 in combination with decreased salinity on the microplanktonic food web. However, long-term effects of the experimental treatments need to be further studied, and indirect effects of the lower salinity treatments could not be ruled out. Our study adds one piece to the complicated puzzle to reveal the combined effects of increased pCO2 and reduced salinity levels on the Baltic microplanktonic community.

Climate change and regulation of hepatotoxin production in Cyanobacteria

Harmful, bloom-forming cyanobacteria (CyanoHABs) are occurring with increasing regularity in freshwater and marine ecosystems. The most commonly occurring cyanobacterial toxins are the hepatotoxic microcystin and nodularin. These cyclic hepta- and pentapeptides are synthesised nonribosomally by the gene products of the toxin gene clusters mcy and nda, respectively. Understanding of the regulation of hepatotoxin production is incomplete, although there is strong evidence supporting the roles of iron, light, higher nitrate availability and inorganic carbon in modulating microcystin levels. The majority of these studies have focused on the unicellular freshwater, microcystin-producing strain of Microcystis aeruginosa, with little attention being paid to terrestrial or marine toxin producers. This review intends to investigate the regulation of microcystin and nodularin production in unicellular and filamentous diazotrophic cyanobacteria against the background of changing climate conditions. Special focus is given to diazotrophic filamentous cyanobacteria, for example Nodularia spumigena, capable of regulating their nitrogen levels by actively fixing dinitrogen. By combining data from significant studies, an overall scheme of the regulation of toxin production is presented, focussing specifically on nodularin production in diazotrophs against the background of increasing carbon dioxide concentrations and temperatures envisaged under current climate change models. Furthermore, the risk of sustaining and spreading CyanoHABs in the future ocean is evaluated.

New Structural Variants of Aeruginosin Produced by the Toxic Bloom Forming Cyanobacterium Nodularia spumigena

Nodularia spumigena is a filamentous diazotrophic cyanobacterium that forms blooms in brackish water bodies. This cyanobacterium produces linear and cyclic peptide protease inhibitors which are thought to be part of a chemical defense against grazers. Here we show that N. spumigena produces structurally novel members of the aeruginosin family of serine protease inhibitors. Extensive chemical analyses including NMR demonstrated that the aeruginosins are comprised of an N-terminal short fatty acid chain, L-Tyr, L-Choi and L-argininal and in some cases pentose sugar. The genome of N. spumigena CCY9414 contains a compact 18-kb aeruginosin gene cluster encoding a peptide synthetase with a reductive release mechanism which offloads the aeruginosins as reactive peptide aldehydes. Analysis of the aeruginosin and spumigin gene clusters revealed two different strategies for the incorporation of N-terminal protecting carboxylic acids. These results demonstrate that strains of N. spumigena produce aeruginosins and spumigins, two families of structurally similar linear peptide aldehydes using separate peptide synthetases. The aeruginosins were chemically diverse and we found 11 structural variants in 16 strains from the Baltic Sea and Australia. Our findings broaden the known structural diversity of the aeruginosin peptide family to include peptides with rare N-terminal short chain (C2–C10) fatty acid moieties.

Insights into the Physiology and Ecology of the Brackish-Water-Adapted Cyanobacterium Nodularia spumigena CCY9414 Based on a Genome-Transcriptome Analysis

Nodularia spumigena is a filamentous diazotrophic cyanobacterium that dominates the annual late summer cyanobacterial blooms in the Baltic Sea. But N. spumigena also is common in brackish water bodies worldwide, suggesting special adaptation allowing it to thrive at moderate salinities. A draft genome analysis of N. spumigena sp. CCY9414 yielded a single scaffold of 5,462,271 nucleotides in length on which genes for 5,294 proteins were annotated. A subsequent strand-specific transcriptome analysis identified more than 6,000 putative transcriptional start sites (TSS). Orphan TSSs located in intergenic regions led us to predict 764 non-coding RNAs, among them 70 copies of a possible retrotransposon and several potential RNA regulators, some of which are also present in other N2-fixing cyanobacteria. Approximately 4% of the total coding capacity is devoted to the production of secondary metabolites, among them the potent hepatotoxin nodularin, the linear spumigin and the cyclic nodulapeptin. The transcriptional complexity associated with genes involved in nitrogen fixation and heterocyst differentiation is considerably smaller compared to other Nostocales. In contrast, sophisticated systems exist for the uptake and assimilation of iron and phosphorus compounds, for the synthesis of compatible solutes, and for the formation of gas vesicles, required for the active control of buoyancy. Hence, the annotation and interpretation of this sequence provides a vast array of clues into the genomic underpinnings of the physiology of this cyanobacterium and indicates in particular a competitive edge of N. spumigena in nutrient-limited brackish water ecosystems.

The freshwater cyanobacterium Anabaena doliolum transformed with ApGSMT-DMT exhibited enhanced salt tolerance and protection to nitrogenase activity, but became halophilic

Glycine betaine (GB) is an important osmolyte synthesized in response to different abiotic stresses, including salinity. The two known pathways of GB synthesis involve: 1) two step oxidation of choline (choline → betaine aldehyde → GB), generally found in plants, microbes and animals; and 2) three step methylation of glycine (glycine → sarcosine → dimethylglycine → GB), mainly found in halophilic archaea, sulphur bacteria and the cyanobacterium Aphanothece (Ap.) halophytica. Here, we transformed a salt-sensitive freshwater diazotrophic filamentous cyanobacterium Anabaena (An.) doliolum with N-methyltransferase genes (ApGSMT-DMT) from Ap. halophytica using the triparental conjugation method. The transformed An. doliolum synthesized and accumulated GB in cells, and showed increased salt tolerance and protection to nitrogenase activity. The salt responsiveness of the transformant was also apparent as GB synthesis increased with increasing concentrations of NaCl in the nutrient solution, and maximal [12.92 µmol (g dry weight)(-1)] in cells growing at 0.5 M NaCl. Therefore, the transformed cyanobacterium has changed its behaviour from preferring freshwater to halophily. This study may have important biotechnological implications for the development of stress tolerant nitrogen-fixing cyanobacteria as biofertilizers for sustainable agriculture.

Genome-wide analysis of putative peroxiredoxin in unicellular and filamentous cyanobacteria.

BackgroundCyanobacteria are photoautotrophic prokaryotes with wide variations in genome sizes and ecological habitats. Peroxiredoxin (PRX) is an important protein that plays essential roles in protecting own cells against reactive oxygen species (ROS). PRXs have been identified from mammals, fungi and higher plants. However, knowledge on cyanobacterial PRXs still remains obscure. With the availability of 37 sequenced cyanobacterial genomes, we performed a comprehensive comparative analysis of PRXs and explored their diversity, distribution, domain structure and evolution.ResultsOverall 244 putative prx genes were identified, which were abundant in filamentous diazotrophic cyanobacteria, Acaryochloris marina MBIC 11017, and unicellular cyanobacteria inhabiting freshwater and hot-springs, while poor in all Prochlorococcus and marine Synechococcus strains. Among these putative genes, 25 open reading frames (ORFs) encoding hypothetical proteins were identified as prx gene family members and the others were already annotated as prx genes. All 244 putative PRXs were classified into five major subfamilies (1-Cys, 2-Cys, BCP, PRX5_like, and PRX-like) according to their domain structures. The catalytic motifs of the cyanobacterial PRXs were similar to those of eukaryotic PRXs and highly conserved in all but the PRX-like subfamily. Classical motif (CXXC) of thioredoxin was detected in protein sequences from the PRX-like subfamily. Phylogenetic tree constructed of catalytic domains coincided well with the domain structures of PRXs and the phylogenies based on 16s rRNA.ConclusionsThe distribution of genes encoding PRXs in different unicellular and filamentous cyanobacteria especially those sub-families like PRX-like or 1-Cys PRX correlate with the genome size, eco-physiology, and physiological properties of the organisms. Cyanobacterial and eukaryotic PRXs share similar conserved motifs, indicating that cyanobacteria adopt similar catalytic mechanisms as eukaryotes. All cyanobacterial PRX proteins share highly similar structures, implying that these genes may originate from a common ancestor. In this study, a general framework of the sequence-structure-function connections of the PRXs was revealed, which may facilitate functional investigations of PRXs in various organisms.

The slow S to M fluorescence rise in cyanobacteria is due to a state 2 to state 1 transition

In dark-adapted plants and algae, chlorophyll a fluorescence induction peaks within 1s after irradiation due to well documented photochemical and non-photochemical processes. Here we show that the much slower fluorescence rise in cyanobacteria (the so-called “S to M rise” in tens of seconds) is due to state 2 to state 1 transition. This has been demonstrated in particular for Synechocystis PCC6803, using its RpaC− mutant (locked in state 1) and its wild-type cells kept in hyperosmotic suspension (locked in state 2). In both cases, the inhibition of state changes correlates with the disappearance of the S to M fluorescence rise, confirming its assignment to the state 2 to state 1 transition. The general physiological relevance of the SM rise is supported by its occurrence in several cyanobacterial strains: Synechococcus (PCC 7942, WH 5701) and diazotrophic single cell cyanobacterium (Cyanothece sp. ATCC 51142). We also show here that the SM fluorescence rise, and also the state transition changes are less prominent in filamentous diazotrophic cyanobacterium Nostoc sp. (PCC 7120) and absent in phycobilisome-less cyanobacterium Prochlorococcus marinus PCC 9511. Surprisingly, it is also absent in the phycobiliprotein rod containing Acaryochloris marina (MBIC 11017). All these results show that the S to M fluorescence rise reflects state 2 to state 1 transition in cyanobacteria with phycobilisomes formed by rods and core parts. We show that the pronounced SM fluorescence rise may reflect a protective mechanism for excess energy dissipation in those cyanobacteria (e.g. in Synechococcus PCC 7942) that are less efficient in other protective mechanisms, such as blue light induced non-photochemical quenching. This article is part of a Special Issue entitled: Photosynthesis Research for Sustainability: from Natural to Artificial.

Interspecific Resource Competition—Combined Effects of Radiation and Nutrient Limitation on Two Diazotrophic Filamentous Cyanobacteria

The cyanobacterial blooms in the Baltic Sea are dominated by diazotrophic cyanobacteria, the potentially toxic species Aphanizomenon sp. and the toxic species Nodularia spumigena. The seasonal succession with peaks of Aphanizomenon sp., followed by peaks of N. spumigena, has been explained by the species-specific niches of the two species. In a three-factorial outdoor experiment, we tested if nutrient and radiation conditions may impact physiological and biochemical responses of N. spumigena and Aphanizomenon sp. in the presence or absence of the other species. The two nutrient treatments were f/2 medium without NO (3) (-) (-N) and f/2 medium without PO (4) (3-) (-P), and the two ambient radiation treatments were photosynthetic active radiation >395nm (PAR) and PAR + UV-A + UV-B >295nm. The study showed that Aphanizomenon sp. was not negatively affected by the presence of N. spumigena and that N. spumigena was better adapted to both N and P limitation in interaction with ultraviolet radiation (UVR, 280-400nm). In the Baltic Sea, these physical conditions are likely to prevail in the surface water during summer. Interestingly, the specific growth rate of N. spumigena was stimulated by the presence of Aphanizomenon sp. We suggest that the seasonal succession, with peaks of Aphanizomenon sp. followed by peaks of N. spumigena, is a result from species-specific preferences of environmental conditions and/or stimulation by Aphanizomenon sp. rather than an allelopathic effect of N. spumigena. The results from our study, together with a predicted stronger stratification due to effects of climate change in the Baltic Sea with increased temperature and increased precipitation and increased UV-B due to ozone losses, reflect a scenario with a continuing future dominance of the toxic N. spumigena.

Does female RNA content reflect viable egg production in copepods? A test with the Baltic copepod Acartia tonsa

During summer, the Baltic Sea is subjected to the world’s largest cyanobacterial blooms. These blooms are linked to eutrophication and raise many questions concerning their effects on the ecosystem. To understand their impacts on the food web dynamics, it is essential to assess growth responses of grazers to these cyanobacteria. In the northern Baltic proper, copepods are the most important herbivores providing an essential link between the primary producers and higher trophic levels. In this Thesis, Papers I & II evaluate methods to estimate copepod growth in response to feeding conditions in situ. The most conspicuous diazotrophic filamentous cyanobacterium in the Baltic Sea is Nodularia spumigena, a producer of nodularin which is highly toxic to vertebrates, yet its ecological role is largely unknown. In Paper III, reciprocal interactions between cyanobacteria, sympatric algae and copepods are studied. The results suggest that nodularin is likely involved in allelopathic interactions, but it is not an inducible defense against grazers. Furthermore, the results of Papers IV & V, indicate that natural assemblages of N. spumigena and Anabaena spp. may support copepod reproduction and that total diazotrophic filamentous cyanobacteria appear to provide a beneficial feeding environment for the feeding stages of copepod nauplii, most probably by stimulating the microbial communities that nauplii feed upon. Since cyanobacterial blooms are projected to increase due to global climate change, the combined effects of toxic cyanobacteria, ocean acidification and global warming predicted for year 2100 are further investigated on copepods in Paper IV. Taken together, these studies indicate that filamentous diazotrophic cyanobacteria contribute to sustaining secondary productivity and have potential implications of management practices with respect to combating eutrophication, global climate change and sustaining fish feeding conditions.

Characterization of Two Critical Residues in the Effector-Binding Domain of NtcA in the Cyanobacterium Anabaena sp. Strain PCC 7120

NtcA, highly conserved in cyanobacteria, regulates the expression of a large number of genes involved in nitrogen and carbon metabolisms, photosynthesis, and stress responses. In the filamentous diazotrophic cyanobacterium Anabaena PCC 7120, NtcA is also required for the initiation of heterocyst differentiation, triggered by the accumulation of 2-oxoglutarate (2-OG) following nitrogen starvation. Recent structural studies reveal the binding pocket of 2-OG on each of the two subunits of the NtcA homodimer, and indicate a route of signal transmission upon 2-OG binding. In this study, we studied the effect of mutations of two critical residues in the effector-binding domain of NtcA on heterocyst differentiation. Mutations of these residues could change strongly the ability of NtcA to sense the nitrogen-starvation signal in vivo. As a result of these mutations, the corresponding strains were unable to form any heterocysts, or form a few heterocysts at a very low frequency. Consistent with these phenotypes, these mutations were defective in initiating transcription by the RNA polymerase in the presence of 2-OG as determined by a reconstituted in vitro transcriptional assay. The different effects of the two mutations were consistent with the roles of the two corresponding residues in 2-OG binding highlighted by recent structural analysis of the NtcA-2-OG complex. These studies provided genetic evidence for the importance of the effector-binding domain in the regulatory function of NtcA.