-
[show abstract]
[hide abstract]
ABSTRACT: Microcystins are potent peptide toxins produced by a range of distantly related cyanobacteria. They are assembled on a large enzyme complex encoded by the 55 kb microcystin synthetase (mcy) gene cluster. Here we report two strains of the genus Anabaena isolated from the Baltic Sea, which contain the entire mcy gene cluster but do not produce microcystins. Transcription analysis demonstrated that mcy genes were not expressed in these strains. We identified short insertion elements interrupting the mcyA, mcyD and mcyE genes in the two strains isolated in different years from different parts of the Baltic Sea. The 126–207 bp insertion elements encoded both terminal inverted repeats and direct repeats but lacked transposases. However, we found evidence for transposition of these elements despite the absence of genes encoding transposases, suggesting that they are non-autonomous mobile miniature inverted-repeat transposable elements (MITEs) recently described from cyanobacteria. The MITE insertion elements were present in mcyD genes amplified directly from the cyanobacterial community present in the Baltic Sea blooms from 2005. The results demonstrate that mcy gene cluster mutants can make up a stable proportion of the mcy gene pool in the Baltic Sea population.
Environmental Microbiology Reports 08/2010; 3(2):189 - 194. · 3.23 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The late summer mass occurrences of cyanobacteria in the Baltic Sea are among the largest in the world. These blooms are rarely monotypic and are often composed of a diverse assemblage of cyanobacteria. The toxicity of the blooms is attributed to Nodularia spumigena through the production of the hepatotoxic nodularin. However, the microcystin hepatotoxins have also been reported from the Baltic Sea on a number of occasions. Recent evidence links microcystin production in the Gulf of Finland directly to the genus Anabaena. Here we developed a denaturing gradient gel electrophoresis (DGGE) method based on the mcyE microcystin synthetase gene and ndaF nodularin synthetase gene that allows the culture-independent discrimination of microcystin- and nodularin-producing cyanobacteria directly from environmental samples. We PCR-amplified microcystin and nodularin synthetase genes from environmental samples taken from the Gulf of Finland and separated them on a denaturing gradient gel using optimized conditions. Sequence analyses demonstrate that uncultured microcystin-producing Anabaena strains are genetically more diverse than previously demonstrated from cultured strains. Furthermore, our data show that microcystin-producing Anabaena are widespread in the open Gulf of Finland. Non-parametric statistical analysis suggested that salinity plays an important role in defining the distribution of microcystin-producing Anabaena. Our results indicate that microcystin-producing blooms are a persistent phenomenon in the Gulf of Finland.
Environmental Microbiology 01/2009; 11(4):855-66. · 5.84 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Late summer cyanobacterial blooms in the Baltic Sea contain Anabaena sp. together with Nodularia spumigena and Aphanizomenon flos-aquae. Although Anabaena is common especially in the Gulf of Finland, very little is known about its genetic diversity. Here we undertook a molecular phylogenetic study of 68 Anabaena strains isolated from the brackish Gulf of Finland. We sequenced the 16S rRNA genes from 54 planktonic and 14 benthic Anabaena strains, and rbcL and rpoC1 genes from a subset of these strains. Phylogenetic trees showed that Anabaena strains, from both planktonic and benthic habitats, were genetically diverse. Although the Anabaena strains were morphologically diverse, in our study only one genetically valid species was found to exist in the plankton. Evolutionary distances between benthic Anabaena strains were greater than between planktonic strains, suggesting that benthic habitats allow for the maintenance of greater genetic diversity than planktonic habitats. A number of novel lineages containing only sequences obtained in this study were compiled in the phylogenetical analyses. Thus, it seemed that novel lineages of the genus Anabaena may be present in the Baltic Sea. Our results demonstrate that the Baltic Sea Anabaena strains show surprisingly high genetic diversity.
FEMS Microbiology Ecology 06/2008; 64(2):199-208. · 3.41 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Anabaena is a filamentous, N(2)-fixing, and morphologically diverse genus of cyanobacteria found in freshwater and brackish water environments worldwide. It contributes to the formation of toxic blooms in freshwater bodies through the production of a range of hepatotoxins or neurotoxins. In the Baltic Sea, Anabaena spp. form late summer blooms, together with Nodularia spumigena and Aphanizomenon flos-aquae. It has been long suspected that Baltic Sea Anabaena may produce microcystins. The presence of microcystins has been reported for the coastal regions of the Baltic proper, and a recent report also indicated the presence of the toxin in the open Gulf of Finland. However, at present there is no direct evidence linking Baltic Sea Anabaena spp. to microcystin production. Here we report on the isolation of microcystin-producing strains of the genus Anabaena in the open Gulf of Finland. The dominant microcystin variants produced by these strains included the highly toxic MCYST-LR as well as [d-Asp(3)]MCYST-LR, [d-Asp(3)]MCYST-HtyR, MCYST-HtyR, [d-Asp(3),Dha(7)]MCYST-HtyR, and [Dha(7)]MCYST-HtyR variants. Toxic strains were isolated from the coastal Gulf of Finland as well as from the easternmost open-sea sampling station, where there were lower salinities than at other stations. This result suggests that lower salinity may favor microcystin-producing Anabaena strains. Furthermore, we sequenced 16S rRNA genes and found evidence for pronounced genetic heterogeneity of the microcystin-producing Anabaena strains. Future studies should take into account the potential presence of microcystin-producing Anabaena sp. in the Gulf of Finland.
Applied and Environmental Microbiology 11/2007; 73(20):6543-50. · 3.83 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: In this article we summarize the current knowledge of Baltic Sea cyanobacteria, focusing on diversity, toxicity, and nitrogen fixation in the filamentous heterocystous taxa. We also review the recent results of our microbial diversity studies in planktonic and benthic habitats in the Baltic Sea. Based on molecular analyses, we have improved the understanding of cyanobacterial population structure by assessing genetic diversity within species that are morphologically inseparable. Moreover, we have studied microbial functions such as toxin production and nitrogen fixation in situ under different environmental conditions. Phosphorus limitation of bloom-forming, nitrogen-fixing cyanobacteria was clearly verified, emphasizing the importance of continuous efforts to reduce this element in the Baltic Sea. We have designed a rapid and reliable detection method for the toxic cyanobacterium Nodularia spumigena, which can be used to study bloom formation of this important toxin producer.
AMBIO A Journal of the Human Environment 05/2007; 36(2-3):180-5. · 2.03 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Cyanobacterial mass occurrences in freshwater lakes are generally formed by Anabaena, Microcystis, and Planktothrix, which may produce cyclic heptapeptide hepatotoxins, microcystins. Thus far, identification of the most potent microcystin producer in a lake has not been possible due to a lack of quantitative methods. The aim of this study was to identify the microcystin-producing genera and to determine the copy numbers of microcystin synthetase gene E (mcyE) in Lake Tuusulanjärvi and Lake Hiidenvesi in Finland by quantitative real-time PCR. The microcystin concentrations and cyanobacterial cell densities of these lakes were also determined. The microcystin concentrations correlated positively with the sum of Microcystis and Anabaena mcyE copy numbers from both Lake Tuusulanjärvi and Lake Hiidenvesi, indicating that mcyE gene copy numbers can be used as surrogates for hepatotoxic Microcystis and ANABAENA: The main microcystin producer in Lake Tuusulanjärvi was Microcystis spp., since average Microcystis mcyE copy numbers were >30 times more abundant than those of ANABAENA: Lake Hiidenvesi seemed to contain both nontoxic and toxic Anabaena as well as toxic Microcystis strains. Identifying the most potent microcystin producer in a lake could be valuable for designing lake restoration strategies, among other uses.
Applied and Environmental Microbiology 12/2003; 69(12):7289-97. · 3.83 Impact Factor
-
Katrianna Halinen
[show abstract]
[hide abstract]
ABSTRACT: Syanobakteerit (sinilevät) ovat olleet Itämeressä koko nykymuotoisen Itämeren ajan, sillä paleolimnologiset todisteet niiden olemassaolosta Itämeren alueella ovat noin 7000 vuoden takaa. Syanobakteerien massaesiintymät eli kukinnat ovat kuitenkin sekä levinneet laajemmille alueille että tulleet voimakkaimmiksi viimeisten vuosikymmenien aikana. Tähän on osasyynä ihmisten aiheuttama kuormitus, joka rehevöittää Itämerta. Suomenlahti, jota tämä tutkimus käsittelee, on kärsinyt tästä rehevöitymiskehityksestä muita Itämeren altaita enemmän. Syanobakteerit muodostavat jokakesäisiä kukintoja Suomenlahdella - niin sen avomerialueilla kuin rannoillakin. Yleisimmät kukintoja muodostavat syanobakteerisuvut ovat Nodularia, Anabaena ja Aphanizomenon. Kukinnat aiheuttavat paitsi esteettistä haittaa myös terveydellisen riskitekijän. Niiden myrkyllisyys liitetään usein Nodularia-suvun tuottamaan nodulariini-maksamyrkkyyn. Itämeren Aphanizomenon-suvun on todettu olevan myrkytön. Vaikka Itämeren kukintoja aiheuttavista Nodularia- ja Aphanizomenon-syanobakteereista tiedetään varsin paljon, on molekyylimenetelmiin pohjautuva syanobakteeritutkimus ohittanut Itämeren Anabaena-suvun monelta osin. Tämän työn tarkoituksena oli syventää käsitystämme Itämeren Anabaena-syanobakteerista, sen mahdollisesta myrkyllisyydestä, geneettisestä monimuotoisuudesta ja fylogeneettisista sukulaisuussuhteista. Tässä työssä eristettiin 49 planktista Anabaena-kantaa, joista viisi tuottivat mikrokystiinejä. Tämä oli ensimmäinen yksiselitteinen todiste, että Itämeren Anabaena tuottaa maksamyrkyllisiä mikrokystiini-yhdisteitä. Jokainen eristetty myrkyllinen Anabaena-kanta tuotti useita mikrokystiini-variantteja. Lisäksi mikrokystiinejä löydettiin kukintanäytteistä, joissa oli myrkkyä syntetisoivia geenejä sisältäneitä Anabaena-syanobakteereita. Myrkkyjä löydettiin molempina tutkimusvuosina 2003 ja 2004. Myrkkyjen esiintyminen ei siten ollut vain yksittäinen ilmiö. Tässä työssä saimme viitteitä siitä, että maksamyrkyllinen Anabaena-syanobakteeri esiintyisi vähäsuolaisissa vesissä. Tämä riippuvuussuhde jää kuitenkin tulevien tutkimuksien selvitettäväksi. Tässä työssä havaittiin mikrokystiinisyntetaasi-geenien inaktivoituminen Itämeren Anabaena-kannassa ja kukintanäytteissä. Kuvasimme Anabaena-kannan mikrokystiinisyntetaasigeenien sisältä insertioita, jotka hyvin todennäköisesti inaktivoivat myrkyntuoton. Insertion sisältäneeltä kannalta löysimme kuitenkin kaikki mikrokystiinisyntetaasigeenit osoittaen, että geenien olemassaolo ei välttämättä varmista kannan mikrokystiinintuottoa. Mielenkiintoista oli se, että inaktivaation aiheuttavia insertioita löytyi kukintanäytteistä molemmilta tutkimusvuosilta. Vastaavia insertioita ei kuitenkaan löydetty makean veden Anabaena-kannoista tai järvinäytteistä. On yleistä, että syanobakteerikukinnoista löytyy usean syanobakteerisuvun edustajia. Myrkyllisiä sukuja tai lajeja ei voida kuitenkaan erottaa mikroskooppisesti myrkyttömistä. Käsillä olevassa tutkimuksessa kehitettiin molekyylimenetelmä, jolla on mahdollista määrittää kukinnan mahdollisesti maksamyrkylliset syanobakteerisuvut. Tätä menetelmää sovellettiin Itämeren kukintojen tutkimiseen. Itämeren pintavesistä ja ranta-alueiden pohjasta eristetyt Anabaena-kannat osoittautuivat geneettisesti monimuotoisiksi. Tämä Anabaena-syanobakteerien geneettinen monimuotoisuus vahvistettiin monistamalla geenejä suoraan kukintanäytteistä ilman kantojen eristystä. Makeiden vesien ja Itämeren Anabaena-kannat ovat geneettisesti hyvin samankaltaisia. Geneettisissä vertailuissa kävi kuitenkin ilmi, että pohjassa elävien Anabaena-kantojen geneettinen monimuotoisuus oli suurempaa kuin pintavesistä eristettyjen kantojen. Itämeren Anabaena-kantojen sekvenssit muodostivat omia ryhmiä sukupuun sisällä, jolloin on mahdollista, että nämä edustavat Itämeren omia Anabaena-ekotyyppejä. Tämä tutkimus oli ensimmäinen, jossa uusin molekyylimenetelmin systemaattisesti selvitettiin Itämeren Anabaena-syanobakteerin geneettistä populaatiorakennetta, fylogeniaa ja myrkyntuottoa. Tulevaisuudessa monitorointitutkimuksissa on otettava huomioon myös Itämeren Anabaena-syanobakteerin mahdollinen maksamyrkyntuotto – erityisesti vähäsuolaisemmilla rannikkovesillä. Cyanobacteria (blue-green algae) form blooms in the Baltic Sea during the warmest summer months. According to paleolimnological data, cyanobacteria have long history in the Baltic Sea, going back at least 7000 years. However, the intensity as well as the expanse of cyanobacterial blooms has increased during recent decades. Blooms attract regular attention in the media because of their visibility and the potential health risk they pose to humans and animals. The Gulf of Finland is the most eutrophied area of the Baltic Sea, and cyanobacterial blooms are widely believed to be the result of intense anthropogenic nutrient loading. Cyanobacterial blooms are formed mainly by species of three genera in the Baltic Sea, Nodularia, Anabaena and Aphanizomenon. The focus of present-day research on Baltic Sea cyanobacteria has been on Nodularia and Aphanizomenon, while the genus Anabaena has been neglected. Anabaena is often considered to play a minor role in cyanobacterial blooms. However, Anabaena can form a significant part of the blooms, especially in the northern part of the Baltic Sea. Cyanobacterial blooms in the Baltic Sea are invariably toxic due to the production of hepatotoxic nodularin by Nodularia spumigena. According to systematic studies, Aphanizomenon flos-aquae was not found to produce hepatotoxins in the Baltic Sea. However, it has been speculated that Baltic Sea Anabaena spp. could produce microcystins. The genetic structure of the Anabaena populations in the Baltic Sea has not been systematically explored. The aim of this present study was to increase our understanding of the Anabaena - a component of the Baltic Sea phytoplankton. Altogether, 49 planktonic Anabaena strains were isolated from the Gulf of Finland, five of which were microcystin-producing. This study provided unequivocal evidence that Baltic Sea Anabaena is able to produce microcystins. Each microcystin-producing Anabaena strain produced two to four dominant microcystin variants, including the highly toxic microcystin-LR. In this study, a culture-independent method was designed to detect putative microcystin and nodularin producers. By means of this DGGE method, microcystin-producing Anabaena populations were detected in cyanobacterial bloom samples from the summers of 2003 and 2004. Microcystin-producing Anabaena populations were detected throughout the Gulf of Finland. This excluded the possibility that the presence of microcystin-producing Anabaena was a chance phenomenon. Results suggest that salinity may limit the distribution of the microcystin-producing Anabaena although further studies are needed to confirm the interdependence of salinity and microcystin production. Microcystin-producing Anabaena populations were found to be highly diverse on analyses of the 16S rRNA, rbcL, rpoC1, and mcyE gene sequences. In previous studies, freshwater microcystin-producing Anabaena strains were grouped together in phylogenetic analyses. All microcystin-producing Baltic Sea Anabaena strains belonged to this hepatotoxic cluster, with the exception of a single strain. Both planktonic and benthic Anabaena populations were genetically heterogeneous and closely related to freshwater Anabaena strains. However, genetic diversity in benthic Anabaena strains was higher than in planktonic strains. In phylogenetic analyses, novel Anabaena lineages, possibly specific to the Baltic Sea, were identified. This suggests ecotypic diversification within Anabaena populations. We found two planktonic Anabaena strains which carried the entire mcy gene cluster, but were nonetheless incapable of producing microcystins. Natural genetic inactivation of the mcy gene cluster was identified in Anabaena strain BIR259. This strain carried insertions which most likely caused the inactivation of the mcy genes. The insertions documented here were surprisingly common in the Baltic Sea bloom samples and they were present in samples from both studied summers, 2003 and 2004. However, these insertions were not identified in freshwater strains or in field samples from freshwater lakes. The aim of this study was to establish a strain collection of Baltic Sea Anabaena and to shed light on the phylogeny, microcystin-production, and genetic diversity of the Baltic Sea Anabaena populations. In addition to strain isolation, these research goals were approached by in situ molecular methods. Systematic toxin screening showed that Anabaena is able to produce microcystins, and this should be taken into account in future toxin monitoring programmes.
