Cyanobacteria For Production Research Articles

An Integrated Approach of Using Polystyrene Foam as an Attachment System for Growth of Mixed Culture of Cyanobacteria with Concomitant Treatment of Copper Mine Waste Water

Cyanobacteria have tremendous applications in areas such as production of biofuels and pharmaceutically important pigments and are used as an adsorbent for the removal of toxic metabolites. However, large scale production of Cyanobacteria is not economically feasible due to high cost involved in separation of biomass. In this context, different attachment systems have been developed for the growth of Cyanobacteria on a solid support. In this study, a simple and economical attachment system using polystyrene foam for growth of Cyanobacteria has been presented. Results clearly indicate that high biomass yield can be obtained in attached system when compared to suspended system. In attachment system, the biomass yield showed 21.4%, total protein content showed 29.2%, chlorophyll content showed 11.1%, and carotenoid content showed 13.1% increase as compared to the suspended system. The attachment system can also support the growth of Cyanobacteria in presence of copper mine waste water with concomitant removal of copper ions. These results were corroborated by COD analysis, which indicated significant reduction. Further, copper removal was high in attached system as compared to suspended system. It appears that attachment system offers protection for growing Cyanobacteria and can be effectively employed for growing Cyanobacteria in presence of waste water coming from different sources.

Phormidium autumnale Growth and Anatoxin-a Production under Iron and Copper Stress

Studies on planktonic cyanobacteria have shown variability in cyanotoxin production, in response to changes in growth phase and environmental factors. Few studies have investigated cyanotoxin regulation in benthic mat-forming species, despite increasing reports on poisoning events caused by ingestion of these organisms. In this study, a method was developed to investigate changes in cyanotoxin quota in liquid cultures of benthic mat-forming cyanobacteria. Iron and copper are important in cellular processes and are well known to affect growth and selected metabolite production in cyanobacteria and algae. The effect of iron (40–4000 μg L−1) and copper (2.5–250 μg L−1) on growth and anatoxin-a quota in Phormidium autumnale was investigated in batch culture. These concentrations were chosen to span those found in freshwater, as well as those previously reported to be toxic to cyanobacteria. Anatoxin-a concentrations varied throughout the growth curve, with a maximum quota of between 0.49 and 0.55 pg cell−1 measured within the first two weeks of growth. Growth rates were significantly affected by copper and iron concentrations (P < 0.0001); however, no statistically significant difference between anatoxin-a quota maxima was observed. When the iron concentrations were 800 and 4000 μg L−1, the P. autumnale cultures did not firmly attach to the substratum. At 250 μg L−1 copper or either 40 or 4000 μg L−1 iron, growth was suppressed.

Discrete Modeling of Dynamics of Zooplankton Community at the Different Stages of an Antropogeneous Eutrophication

Mathematical modeling is a convenient way for characterization of complex ecosystems. This approach was applied to study the dynamics of zooplankton in Lake Sevan (Armenia) at different stages of anthropogenic eutrophication with the use of a novel method called discrete modeling of dynamical systems with feedback (DMDS). Simulation demonstrated that the application of this method helps in characterization of inter- and intra-component relationships in a natural ecosystem. This method describes all possible pairwise inter-component relationships like "plus-plus," "minus-minus," "plus-minus," "plus-zero," "minus-zero," and "zero-zero" that occur in most ecosystems. Based on the results, a working hypothesis was formulated. It was found that the sensitivity to weak external influence in zooplanktons was the greatest during the mid period of eutrophication in Lake Sevan, whereas in the final stages of eutrophication, an outbreak in the biomass production of cyanobacteria was evident. To support this approach, a weak external disturbance in the form of magnetic storm was used to see its effect on species Daphnia longispina sevanica. A statistically significant correlation between the frequency of magnetic storms and the number of this species was revealed and an increase in the number of toxic cyanobacteria species as a consequence of eutrophication. This paper, for the first time, suggests a DMDS method, to diagnose impact of anthropogenic eutrophication on environment.

Does the presence of zooplankton influence growth and toxin production ofNodularia spumigena?

The ecological role of toxin production in cyanobacteria is still unknown. Here, we studied the potential effect of different zooplankton taxa and density on growth and toxin production of the cyanobacterium Nodularia spumigena. In this experiment, N. spumigena was exposed to three different zooplankton taxa: Rotifera (Synchaeta baltica), Copepoda (Eurytemora affinis) and Cladocera (Daphnia magna) and as a comparison a zooplankton free control. We used false bottom chambers separating the cyanobacterium and zooplankton and were thereby able to prevent direct grazing. Overall, N. spumigena toxin levels (nodularin expressed as microcystin equivalents) were significantly lower when exposed to the different zooplankton compared to the zooplankton free control, whereas no effects could be detected on N. spumigena growth and biomass. Although we did not test the underlying mechanism, these findings suggest that the presence of zooplankton reduce toxin production in N. spumigena, independent of growth. Therefore, toxin production in N. spumigena should not be considered an induced defence mechanism against zooplankton, at least not on a short-term basis.

Hydrocarbon profiles and phylogenetic analyses of diversified cyanobacterial species

The combination of environmental concerns and the growing demand for energy make the development of biofuels, an attractive alternative to fossil fuels, a goal for many researchers. The direct photosynthetic production of hydrocarbons, which are the major components of fossil fuels, is considered to be a promising and innovative strategy for the development of biofuels with advanced fuel properties and solar-driven energy input. Cyanobacteria have existed continuously since the early evolution of the biosphere and the biosynthetic pathways of hydrocarbons in these prokaryotes have been genetically and biochemically identified. In this study, the hydrocarbon compositions of 19 freshwater cyanobacterial species distributed among 13 genera were analyzed. Based on their hydrocarbon profiles, these cyanobacterial species were classified into 5 major subgroups. Combined with the previously reported hydrocarbon compositions in different cyanobacterial species, we found that branched-chain alkanes were limited predominantly in filamentous species but rarely in unicellular species. Phylogenetic analysis using traditional small-subunit ribosomal RNA (16S rDNA) of these strains presented clustering similar to their hydrocarbon production profiles. Acyl–acyl carrier protein reductase (AAR) and aldehyde deformylating oxygenase (ADO) are two key enzymes involved in the biosynthesis of hydrocarbons in cyanobacteria. A comparison of phylogenies revealed that the topology of 16S rDNA showed a general congruence with that of AAR but not with that of ADO. The results not only provide an evolutionary perspective with which to study the physiological function of cellular hydrocarbons but also display the engineering capacity to molecularly design diversified hydrocarbon fuel products in cyanobacteria.

Combustion and emissions characterization of terpenes with a view to their biological production in cyanobacteria

In developing future fuels there is an opportunity to make use of advances in many fields of science and engineering to ensure that such fuels are sustainable in both production and utilization. One such advance is the use of synthetic biology to re-engineer photosynthetic micro-organisms such that they are able to produce novel hydrocarbons directly from CO2. Terpenes are a class of hydrocarbons that can be produced biologically and have potential as liquid transport fuels. This paper presents experimental studies on a compression ignition engine and spark ignition engine in which the combustion and emissions of 12 different terpenes that could potentially be produced by cyanobacteria were assessed as single components and blends with fossil diesel and fossil gasoline. The 12 terpenes were chosen to explore how small changes to the molecular structure of geraniol (a terpene most easily produced by cyanobacteria) impact on combustion and emissions. Furthermore, the toxicity of some of the best performing terpenes were assessed using the model cyanobacterium Synechocystis sp. PCC6803 (hereafter, Synechocystis) as a prelude to a metabolic engineering programme. The compression ignition engine tests were carried out at constant injection timing and constant ignition timing, and the spark ignition engine tests were conducted at a constant spark timing and a constant lambda value of 1. Of the terpenes tested in the compression ignition engine, geranial and farnesene were found to be the best performing single component fuels in terms of combustion and emissions. In blends with fossil diesel, the presence of geranial or farnesene did not have a significant effect on combustion phasing up to a terpene content of 20% (wt/wt), though levels of NOx and CO did increase. In the spark ignition engine experiments of terpene and fossil gasoline blends, citronellene and linalool were found to be soluble in fossil gasoline and combusted in a steady manner up to a terpene content of 45% and 65% (wt/wt) respectively. Of those terpenes with the most potential as either diesel or gasoline fuels, geraniol and geranial were found to be the most toxic to Synechocystis, with farnesene and linalool less toxic and citronellene having no detrimental effect. Addition of n-dodecane to the cultures was found to ameliorate the toxic effects of all five terpenes.

Engineering cyanobacteria to improve photosynthetic production of alka(e)nes

BackgroundCyanobacteria can utilize solar energy and convert carbon dioxide into biofuel molecules in one single biological system. Synechocystis sp. PCC 6803 is a model cyanobacterium for basic and applied research. Alkanes are the major constituents of gasoline, diesel and jet fuels. A two-step alkane biosynthetic pathway was identified in cyanobacteria recently. It opens a door to achieve photosynthetic production of alka(e)nes with high efficiency by genetically engineering cyanobacteria.ResultsA series of Synechocystis sp. PCC6803 mutant strains have been constructed and confirmed. Overexpression of both acyl-acyl carrier protein reductase and aldehyde-deformylating oxygenase from several cyanobacteria strains led to a doubled alka(e)ne production. Redirecting the carbon flux to acyl- ACP can provide larger precursor pool for further conversion to alka(e)nes. In combination with the overexpression of alkane biosynthetic genes, alka(e)ne production was significantly improved in these engineered strains. Alka(e)ne content in a Synechocystis mutant harboring alkane biosynthetic genes over-expressed in both slr0168 and slr1556 gene loci (LX56) was 1.3% of cell dry weight, which was enhanced by 8.3 times compared with wildtype strain (0.14% of cell dry weight) cultivated in shake flasks. Both LX56 mutant and the wildtype strain were cultivated in column photo-bioreactors, and the alka(e)ne production in LX56 mutant was 26 mg/L (1.1% of cell dry weight), which was enhanced by 8 times compared with wildtype strain (0.13% of cell dry weight).ConclusionsThe extent of alka(e)ne production could correlate positively with the expression level of alkane biosynthetic genes. Redirecting the carbon flux to acyl-ACP and overexpressing alkane biosynthetic genes simultaneously can enhance alka(e)ne production in cyanobacteria effectively.

Potent toxins in Arctic environments – Presence of saxitoxins and an unusual microcystin variant in Arctic freshwater ecosystems

Cyanobacteria are the predominant phototrophs in freshwater ecosystems of the polar regions where they commonly form extensive benthic mats. Despite their major biological role in these ecosystems, little attention has been paid to their physiology and biochemistry. An important feature of cyanobacteria from the temperate and tropical regions is the production of a large variety of toxic secondary metabolites. In Antarctica, and more recently in the Arctic, the cyanobacterial toxins microcystin and nodularin (Antarctic only) have been detected in freshwater microbial mats. To date other cyanobacterial toxins have not been reported from these locations. Five Arctic cyanobacterial communities were screened for saxitoxin, another common cyanobacterial toxin, and microcystins using immunological, spectroscopic and molecular methods. Saxitoxin was detected for the first time in cyanobacteria from the Arctic. In addition, an unusual microcystin variant was identified using liquid chromatography–mass spectrometry. Gene expression analyses confirmed the analytical findings, whereby parts of the sxt and mcy operon involved in saxitoxin and microcystin synthesis, were detected and sequenced in one and five of the Arctic cyanobacterial samples, respectively. The detection of these compounds in the cryosphere improves the understanding of the biogeography and distribution of toxic cyanobacteria globally. The sequences of sxt and mcy genes provided from this habitat for the first time may help to clarify the evolutionary origin of toxin production in cyanobacteria.

Cyanobacterial conversion of carbon dioxide to 2,3-butanediol

Conversion of CO(2) for the synthesis of chemicals by photosynthetic organisms is an attractive target for establishing independence from fossil reserves. However, synthetic pathway construction in cyanobacteria is still in its infancy compared with model fermentative organisms. Here we systematically developed the 2,3-butanediol (23BD) biosynthetic pathway in Synechococcus elongatus PCC7942 as a model system to establish design methods for efficient exogenous chemical production in cyanobacteria. We identified 23BD as a target chemical with low host toxicity, and designed an oxygen-insensitive, cofactor-matched biosynthetic pathway coupled with irreversible enzymatic steps to create a driving force toward the target. Production of 23BD from CO(2) reached 2.38 g/L, which is a significant increase for chemical production from exogenous pathways in cyanobacteria. This work demonstrates that developing strong design methods can continue to increase chemical production in cyanobacteria.

Reprogramming the glycolytic pathway for increased hydrogen production in cyanobacteria: metabolic engineering of NAD+-dependent GAPDH

Catabolism of glycogen stored by cyanobacteria occurs during anaerobic auto-fermentation and produces a range of C1–C3 fermentation products and hydrogen via hydrogenase. We investigated both augmenting and rerouting this carbon catabolism by engineering the glycolysis pathway at the NAD+-dependent glyceraldehyde-3-phosphate dehydrogenase (GAPDH-1), its major regulation site at the nexus of two pathways (Oxidative Pentose Phosphate pathway, OPP, and glycolysis/gluconeogenesis). Null (gap1::aphII) and overexpression (gap1+) strains of Synechococcus sp. strain PCC 7002 were constructed in order to produce more NADPH (via rerouting carbon through OPP) and more NADH (via opening the glycolytic bottleneck), respectively. For gap1::aphII quantitative analyses after four days of dark auto-fermentation showed undiminished glycogen catabolism rate, significant increases of intracellular metabolites in both OPP and upper-glycolysis, decrease in lower-glycolysis intermediates, 5.7-fold increase in NADPH, 2.3-fold increase in hydrogen and 1.25-fold increase in CO2vs. wild type (WT). These changes demonstrate the expected outcome of redirection of carbon catabolism through the OPP pathway with significant stimulation of OPP product yields. The gap1+ strain exhibits a large 17% increase in accumulation of glycogen during the prior photoautotrophic growth stage (gluconeogenesis), in parallel with a 2-fold increase in the total [NAD+ + NADH] pool, foreshadowing an increased catabolic capacity. Indeed, the rate of glycogen catabolism during subsequent dark auto-fermentation increased significantly (58%) vs. WT, resulting in increases in both NADH (4.0-fold) and NADPH (2.9-fold) pools, and terminal fermentation products, hydrogen (3.0-fold) D-lactate (2.3-fold) and acetate (1.4-fold). The overall energy conversion yield over four days from catabolized glycogen to hydrogen increased from 0.6 mole of hydrogen per mole of glucose (WT) to 1.4 (gap1::aphII) and 1.1 (gap1+) under headspace accumulation conditions (without hydrogen milking). These findings demonstrate the significant potential of metabolic engineering for redirecting carbon pathways for carbohydrate catabolism and hydrogen production in cyanobacteria.

Temperature-sensitive PSII and promiscuous PSI as a possible solution for sustainable photosynthetic hydrogen production

Sustainable hydrogen production in cyanobacteria becomes feasible as a result of our recent studies of the structure of photosystem I encoding operon in a marine phage. We demonstrated that the fused PsaJF subunit from the phage, substituted for the two separate subunits in Synechocystis, enabled the mutated PSI to accept electrons from additional electron donors such as respiratory cytochromes. In this way, a type of photorespiration was created in which the cell consumes organic material through respiratory processes and PSI serves as a terminal electron acceptor, substituting for cytochrome oxidase. We designed a hydrogen-producing bioreactor in which this type of photorespiration could utilize the organic material of the cell as an electron source for H2 production. We propose, in parallel, to engineer cyanobacterial and/or algal strains with a temperature-sensitive PSII and enhanced respiration rates to achieve efficient and sustainable hydrogen production. This article is part of a Special Issue entitled: Photosynthesis Research for Sustainability: from Natural to Artificial.

The Structure of an Aldehyde Decarbonylase from Prochlorococcus Marinus Implicated in Biofuel Synthesis Indicates an Octadecanoic Acid Ligand Coordinated to a Di-Iron Catalytic Center∗

Prochlorococcus marinus, a prevalent marine cyanobacterium, can use sunlight and atmospheric CO2 to convert fatty-acid like precursors into alkane biofuels. Schirmer et al. (Science, 2010) showed that alkane production in cyanobacteria entails a two-enzyme process in which the acyl-CoA moiety is reduced to an aldehyde followed by decarbonylation to an alkane. The structure of the aldehyde decarbonylase has been determined by the Joint Center for Structural Genomics (PDB ID 2oc5). Here, we have identified the bound metals at the catalytic site as Fe and, based on the results of crystallographic refinement, have assigned a previously unidentified ligand as an octadecanoic acid. The metal identity was determined from X-ray fluorescence and anomalous difference Fourier maps. The oxygen atoms of the octadecanoic acid are within coordination distance of the Fe atoms and the hydrocarbon tail extends into the interior of the protein. Previous studies showed the enzyme can use octadecanal as a substrate, but the structure raises the question if it can directly catalyze alkane formation from octadecanoic acid ∗Supported by NIGMS PSI grants U54 GM094586 and GM074898View Large Image | View Hi-Res Image | Download PowerPoint Slide

THE TOXIC CYANOBACTERIUM NOSTOC SP. STRAIN 152 PRODUCES HIGHEST AMOUNTS OF MICROCYSTIN AND NOSTOPHYCIN UNDER STRESS CONDITIONS.

The understanding of how environmental factors regulate toxic secondary metabolite production in cyanobacteria is important to guarantee water quality. Very little is known on the regulation of toxic secondary metabolite production in benthic cyanobacteria. In this study the physiological regulation of the production of the toxic heptapeptide microcystin (MC) and the non-toxic related peptide nostophycin (NP) in the benthic cyanobacterium Nostoc sp. strain 152 was studied under contrasting environmental conditions. I used a 2(k) levels factorial design, where k is the number of four factors that have been tested: Reduction in temperature (20 vs. 12°C), irradiance (50 vs. 1 μmol · m(-2) · s(-1)), P-PO(4) (144 vs. 0.14 μM P-PO(4)), N-NO(3) (5.88 mM vs. N-NO(3) free). While the growth rate was reduced more than hundred fold under most severe conditions of temperature, irradiance, and phosphate reduction the production of MC and NP never ceased. The MC and NP contents per cell varied at maximum 5- and 10.6-fold each, however the physiological variation did not outweigh the highly significant linear relationship between the daily cell division rate and the MC and NP net production rates. Surprisingly the MC and NP contents per cell showed a maximum under P-PO(4) reduced and irradiance reduced conditions. Both intra- and extracellular MC and NP concentrations were negatively related to P-PO(4) and irradiance. It is concluded that the proximate factor behind maximal cellular MC and NP contents is physiological stress.

Cadmium(II) sequestration characteristics by two isolates of Synechocystis sp. in terms of exopolysaccharide (EPS) production and monomer composition

We investigated cadmium(II) resistance and its association with exopolysaccharide (EPS) production in cyanobacteria. Increased EPS production was associated with Cd(II) resistance. The most resistant isolate, Synechocystis sp. BASO670, secreted the greatest amount of EPS (548 mg/L). EPS production by Synechocystis sp. BASO670 and Synechocystis sp. BASO672 was increased following exposure to 15 and 35 ppm Cd(II). Monomer composition of EPS belonging to each isolate was changed after Cd(II) treatment. Uronic acid contents of Cd(II) treated cells were higher than control cells of each isolate. Scanning electron microscopy (SEM) and energy dispersive X-ray spectroscopy (EDS) analysis confirmed that a considerable amount of metals had precipitated on the cell surface. Fourier transform infrared (FT-IR) spectrum analysis of EPSs belonging to both isolates indicated the presence of C-H and C-O group, which may serve as binding sites for divalent cations.

Phycocyanin promoter of Spirulina platensis controlling heterologous expression in cyanobacteria

An efficient expression vector for transferring exogenous deoxyribonucleic acid (DNA) into cell is important for biological study and recombinant protein production in cyanobacteria. The promoter, which can mediate gene expression and function in both Escherichia coli and cyanobacteria, could facilitate development of an expression vector for a DNA transfer system in cyanobacteria. Computational analysis of the phycocyanin (PC) promoter of Spirulina platensis strain C1 showed a high degree of conservation with the PC promoter of other cyanobacteria. In addition, the S. platensis PC promoter matched well with the consensus sequences of the E. coli promoter. Using green fluorescent protein (GFP) as a reporter, activity of the PC promoter of S. platensis C1 was functionally analyzed in Synechococcus elongatus and E. coli. The result revealed that the PC promoter could efficiently drive gfp expression in heterologous hosts, indicating the existence of common transcription machinery in these microbes. This promoter may be applicable not only for fundamental research but also for biotechnological purposes.

Modulation of IAA Production in Cyanobacteria by Tryptophan and Light

Cyanobacteria represent less a investigated group of prokaryote, in terms of their plant growth promoting potential, especially in relation to the production of phytohormones. The present investigation was aimed towards analyzing growth kinetics, indole acetic acid (IAA) production and acetylene reduction activity (ARA) as an index of nitrogen fixation in two selected cyanobacterial strains belonging to the genus Anabaena, as influenced by tryptophan supplementation and light:dark conditions. Interesting observations were recorded in terms of enhancement of IAA production accompanied by protein and chlorophyll accumulation in the two cyanobacterial strains grown in media without tryptophan and incubated under light:dark or continuous light conditions. Colorimetric and chromatographic analyses supported the observations that tryptophan is not essential as a precursor for IAA biosynthesis in these cyanobacteria. Further study is in progress to identify genes involved in the tryptophan independent pathway for IAA biosynthesis.

Engineering a platform for photosynthetic isoprene production in cyanobacteria, using Synechocystis as the model organism

The concept of “photosynthetic biofuels” envisions application of a single organism, acting both as photo-catalyst and producer of ready-made fuel. This concept was applied upon genetic engineering of the cyanobacterium Synechocystis, conferring the ability to generate volatile isoprene hydrocarbons from CO 2 and H 2O. Heterologous expression of the Pueraria montana (kudzu) isoprene synthase ( IspS) gene in Synechocystis enabled photosynthetic isoprene generation in these cyanobacteria. Codon-use optimization of the kudzu IspS gene improved expression of the isoprene synthase in Synechocystis. Use of the photosynthesis psbA2 promoter, to drive the expression of the IspS gene , resulted in a light-intensity-dependent isoprene synthase expression. Results showed that oxygenic photosynthesis can be re-directed to generate useful small volatile hydrocarbons, while consuming CO 2, without a prior requirement for the harvesting, dewatering and processing of the respective biomass.

Modification of exopolysaccharide composition and production by three cyanobacterial isolates under salt stress

Polysaccharides are renewable resources representing an important class of polymeric materials of biotechnological interest, offering a wide variety of potentially useful products to mankind. Exopolysaccharides (EPSs) of microbial origin with a novel functionality, reproducible physico-chemical properties, stable cost and supply, became a better alternative to polysaccharides of algal origin. EPSs are believed to protect bacterial cells from desiccation, heavy metals or other environmental stresses, including hostimmune responses, and to produce biofilms, thus enhancing the cells chances of colonising special ecological niches. One of the most important stress factor is salt stress for microorganisms. The present investigation is aimed to determine correlation between salt resistance and EPS production by three cyanobacterial isolates (Synechocystis sp. BASO444, Synechocystis sp. BASO507 and Synechocystis sp. BASO511). It is also aimed to investigate the effect of salt concentrations on EPS production by cyanobacteria and effect of salt on monosaccharide composition of EPS. Cyanobacterial isolates were identified by 16 S rRNA analysis. Its salt (NaCl) tolerance and association with exopolysaccharides (EPSs) production in three cyanobacterial isolates were investigated. Also, EPS was analysed by HPLC for monomer characterization. Increased EPS production was associated with NaCl tolerance. The most tolerant isolate, Synechocystis sp. BASO444, secreted the most EPS (500 mg/L). EPS production by Synechocystis sp. BASO444, Synechocystis sp. BASO507 and Synechocystis sp. BASO511 was investigated following exposure to 0.2 and 0.4 M NaCl. Also, flasks containing medium without NaCl were inoculated in the same manner to serve as controls. The monosaccharide compositions of EPS produced by the three isolates following exposure to 0.2 M NaCl were analysed by HPLC. Control EPS of BASO444 was composed of glucose (97%) and galacturonic acid (3%). The composition of BASO511 (control) was glucose (95%), xylose (4.80%), arabinose (0.13%), glucuronic acid (0.03%) and galacturonic acid (0.04%). However, the composition of BASO507 (control) was glucose (0.98%), xylose (98.00%), arabinose (1.00%), glucuronic acid (0.01%) and galacturonic acid (0.01%). In the presence of 0.2 M NaCl, EPS compositions and ratios of three cyanobacterial isolates changed. Although hyperproduction of EPS in response to starvation, antiviral activity, thickening agent and cosmetic industry for product formulations has been reported for cyanobacteria, the effect of NaCl on EPS production in cyanobacteria is not a popular area of study. There are no clear reports correlating EPS production and NaCl tolerance. The gap in the data about the effect of NaCl on cyanobacterial EPS production was filled by this investigation, and the results of our study have important implications in both the industrial and environmental arenas. Our results indicate that 1) exposure to elevated concentrations of NaCl affects the composition of EPS produced by Synechocystis sp. BASO444, Synechocystis sp. BASO507 and Synechocystis sp. BASO511, and 2) there is a correlation between NaCl tolerance and EPS production in some cyanobacteria. Differences in the monosaccharide composition and ratios of EPS may promote NaCl tolerance in these microorganisms. As well, these alternative composition polysaccharides may be important for industrial applications.

The Genetic Basis of Toxin Production in Cyanobacteria

The increasing incidence of mass developments of Cyanobacteria in fresh- and brackish water is a matter of growing concern due to the production of toxins that threaten human and livestock health. The toxins that are produced by freshwater Cyanobacteria comprise hepatotoxins (cyclic peptides such as microcystins and nodularin, as well as alkaloids such as cylindrospermopsin) and neurotoxins (alkaloids such as anatoxin-a, anatoxin-a(S) and saxitoxins). The variation in toxicity between and within species of Cyanobacteria has been recognised for a long time. However, the toxic and non-toxic genotypes within a species cannot be discriminated under the microscope, which has been a major obstacle in identifying those factors that influence toxin production both in the laboratory and in the field. During the last decade, major advances were achieved due to the elucidation and functional characterisation of genes, such as the gene cluster encoding the synthesis of the hepatotoxic heptapeptide, microcystin. Genetic techniques, in particular, have been used to explore (i) the genetic basis, biosynthesis pathways, and physiological regulation of toxin (microcystin) production, (ii) gene loss processes resulting in a patchy distribution of the microcystin synthetase gene cluster among genera and species, as well as (iii) the distribution and abundance of the microcystin genes in the environment. In recent years, experience in detecting microcystin genes directly in the field has increased enormously and robust protocols for the extraction of DNA and the subsequent detection of genes by PCR (polymerase chain reaction)based methods are now available. Due to the high sensitivity of PCR, it is possible to detect toxic genotypes long before a toxic cyanobacterial bloom may occur. Consequently, waterbodies that are at risk of toxic bloom formation can be identified early on in the growing season along with environmental factors that can potentially influence the abundance of toxin producing genotypes.

Isolation and Characterization of the Gene Associated with Geosmin Production in Cyanobacteria

Geosmin is a secondary metabolite responsible for earthy tastes and odors in potable water supplies. Geosmin continues to be a challenge to water utility management regimes and remains one of the most common causes of consumer complaints, as the taste of "dirty" water may suggest a failed disinfection regime and that the water may be unsafe to drink. Although cyanobacteria have been reported to be largely responsible for these taste and odor events, the answer as to how or why geosmin is produced has eluded researchers. We describe here for the first time the mechanism by which geosmin is produced in a model cyanobacterium, Nostoc punctiforme PCC 73102 (ATCC 29133), which we demonstrate utilizes a single enzyme to catalyze the cyclization of farnesyl diphosphate to geosmin. Using this information, we have developed a PCR-based assay that allows the rapid detection of geosmin-producing cyanobacteria. This test may be utilized to confirm and track the emergence of taste and odor-producing cyanobacteria in any given water body and thus can be used as an early warning system by managers of water bodies that may suffer from adverse taste and odor episodes.